CYANONEWS
Volume 11 Number 2 July 1995
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CYANONEWS - a newsletter intended to provide cyanobacteriologists with a forum
for rapid informal communication, unavailable through journals.
Everything you read in this newsletter is contributed by readers like
yourself. Published occasionally, about three times per year.
SUBSCRIPTIONS - $10 or equivalent/year. (See address label for expiration
date). No charge for electronic version.
CONTRIBUTIONS - Expected every couple of years: a new result, an upcoming
meeting or a summary of a past meeting, a post-doctoral opening, a new
publication, a request for strains, a change of life... something. See
last page for addresses you can send news to.
HOW TO FIND OUT MORE ABOUT SOMETHING YOU READ HERE - Each news item contains,
prominently displayed, the name of a contact person. A Directory of
Cyanobacteriologists is distributed every two years or on request.
INSTRUCTIONS TO AUTHORS - Send news.
COPYRIGHT - This newsletter is not copyrighted and no rights are reserved. You
are encouraged to reproduce or to transmit any part of this publication
by whatever means at your disposal, no permission required.
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CONTENTS*CONTENTS*CONTENTS*CONTENTS*CONTENTS*CONTENTS*CONTENTS*CONTENTS*CONTEN
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BULLETIN BOARD
* Collaborators sought for study of photosystem segregation and stacking
* Spirulina biotech offer
* "Toxic Microcystis" published
* Request for news, comments on pigment proteins for review
* Meetings
* Positions sought, Positions available
TRANSITIONS
* Comings and goings of ourselves
* David Laudenbach
NEWS
* Allen's teaching toasted
* Foreign gene expression tamed
* Immobilized cells' boosted NH3 output tied to glutamine synthetase
expression
* Meeting Report: Congress on N2 Fixation
* Meeting report: Euro Workshop on Mol Bio
REFERENCES
ADDRESSES
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BULLETIN BOARD*BULLETIN BOARD*BULLETIN BOARD*BULLETIN BOARD*BULLETIN BOARD*BUL
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****** Matters Arising ******
Dalibor Stys seeks to complement his physico-chemical expertise and resources
with genetically- or theoretically-minded COLLABORATORS to study the influence
of individual proteins from thylakoid proteins on ion and temperature induced
CHANGES IN PHOTOSYSTEM SEGREGATION AND STACKING. He wants to use thylakoid
membranes for studies on specific and unspecific lipid-protein and
protein-protein interactions as well as formation of membrane domains and
ion-induced interactions between membrane lamellae.
He has access to and experience with many spectroscopical techniques and is
looking for collaboration with any group that will supply him with mutants
with defined modifications of thylakoid membranes.
CONTACT: Dalibor Stys, Plant Cell Biology, Box 7007, 220 07 Lund, Sweden.
Tel: 46-46-222-3318, Fax: 46-46-222-4009,
E-mail: Placebio-Dali@Macpost.Lu.Se or Dalibor.Stys@Placebio.Lu.Se
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L.V. Venkataraman offers to provide SPIRULINA TECHNOLOGY, on all aspects
regarding setting up plants of various capacities, including information on
product formulations.
CONTACT: L.V. Venkataraman, "Sudarshana", #236, 8th Cross, Gokulam 3rd
Stage, Mysore 570 002 INDIA. TEL: 821-510006, FAX: 821-512539,
TELEX: 0846-320 POLY IN
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CRC Press has just published a new monograph entitled TOXIC MICROCYSTIS,
edited by M.F. Watanabe, K.I. Harada, W.W. Carmichael, and H. Fujiki. It
includes chapters on the ecology of Microcystis, and the chemistry and
biologically effects of its toxins. The book is 400 pages long and costs
US$189.95 (within U.S.A.) and US$228 (outside U.S.A.)
CONTACT: CRC Press, 2000 Corporate Blvd., N.W., Boca Raton, FL 33431-9868
U.S.A. TEL: 800-272-7737 (within U.S.A.) 407-994-0555 (outside U.S.A.)
FAX: 800-374-3401 (within U.S.A.)
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Beverley Green is writing a REVIEW ON PIGMENT-PROTEINS for Annual Review of
Plant Physiology, to be published in 1996 and would be grateful reprints of
articles she may not have caught. She would also appreciate comments or
questions on controversial points or any other aspect. The review covers
cyanobacterial as well as plant proteins, structure determination,
macromolecular organization, and molecular evolution.
CONTACT: Beverley R. Green, Botany Dept., University of B.C., Vancouver,
B.C. V6T 1Z4 CANADA. TEL: 604-822-2349, FAX: 604-822-6089,
E-MAIL: Beverley.Green@Mtsg.Ubc.Ca
****** MEETINGS ******
The 15TH NORTH AMERICAN SYMBIOTIC NITROGEN FIXATION CONFERENCE will be held
13-18 August 1995 at North Carolina State University, Raleigh.
CONTACT: Gerald Elkan, Department of Microbiology, North Carolina State
University, Box 7615, Raleigh, NC 27695-7615 U.S.A., TEL: 919-515-3945
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A forum on the BIOTECHNOLOGY OF ALGAE will be held in Lake San Marcos Resort,
San Diego, California (U.S.A.) 20 Sept 1995 in conjunction with the
International Symposium on Plant DNA Preservation (17-20 Sept 1995).
CONTACT: E-MAIL: jonthn@aol.com
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The FIRST INTERNATIONAL CONGRESS ON TOXIC CYANOBACTERIA is the new descendent
of the formerly biannual Nordic Symposia on Toxin-producing Algae. The
Congress will be held on the Danish island of Bornholm in the Baltic on 20-24
August 1995. It is planned that the proceedings will be published.
CONTACT: Peter Henriksen, Dept. of Phycology, Botanical Institute,
OE. Farimagsgade 2 D, DK-1353 Copenhagen K, DENMARK. TEL: 45-35-32-22-90
or 45-35-32-22-99, FAX: 45-35-32-23-21, E-MAIL: PHenriks@Bot.Ku.Dk
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The INTERNATIONAL ASSOCIATION OF APPLIED ALGOLOGY will hold its Congress in
South Africa from 16-19 April 1996. Topics include algal production systems,
photosynthesis and physiology, waste water treatment, and commercial ventures.
Registration by the deadline of 30 Nov 1995 is US$200.
CONTACT: Johan Grobbelaar, Bloemfontein, Department of Botany and Genetics,
University of the OFS, Bloemfontein 9300, SOUTH AFRICA. TEL: 27-51-
4012514, FAX: 27-51-488772, E-MAIL: pjg@Rs.Uovs.Ac.Za
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The 13TH INTERNATIONAL SYMPOSIUM ON CYANOPHYTE RESEARCH will take place in
Rome 27 Aug to 3 Sep 1995. The Symposium will focus on taxonomy, extreme
environments, biodiversity, cyanobacterial associations with other organisms,
and ecophysiology. Registration is 200,000 lira. Meals and hotel
accommodations start at 900,000 lira for the nine day symposium.
CONTACT: Patrizia Albertano, Department of Biology, University of Rome `Tor
Vergata', via della Ricerca scientifica, 00133 Rome Italy.
TEL: 39-6-72594345, FAX: 39-6-2023500,
E-MAIL: Albertano@Tovvx1.Ccd.Utovrm.It
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The EUROPEAN SOCIETY FOR PHOTOBIOLOGY will hold its 6th Congress in Cambridge
(Churchill College) from 2nd to 9th September 1995. The congress will have
special session on "Carotenoids in Photosynthesis and Medicine" and "Appli-
cation of protein engineering for the study of light reactions of oxygenic
photosynthesis"
CONTACT: Paul Heelis, Faculty of Science, Health and Medical Studies, The
North East Wales Institute, Plas Coch, Mold Road, Wrexham, Clwyd, LLI
2AW,UK. FAX: 44 (0) 1978 290008, E-MAIL: Heelisp@Newi.Ac.Uk
****** POSITIONS OFFERED ******
POSITION OFFERED: Post-Doc
CONTACT: C.A. Rebeiz, Laboratory of Plant Pigment Biochemistry and
Photobiology, 240 A, PABL, 1201 West Gregory Avenue, University of
Illinois, Urbana IL 61801 U.S.A. TEL: 217-333-1968,
E-MAIL: Tino@Vmd.Cso.Uiuc.Edu
RESEARCH: Either: (1) Study of apoprotein-chlorophyll interaction during the
biosynthesis and assembly of functional light harvesting Chl a/b protein
(LHC II) in higher plants, or (2) Cloning the [4-vinyl]chlorophyllide
a reductase (4VCR) gene [Biochemistry 31:8460-8464 (1992)], an enzyme
responsible for the heterogeneity of chlorophyll biosynthesis in plants
[Ciba Foundation symposium 180, p177-193 (1994)].
REQUIREMENTS: Some expertise in one or more of the following: porphyrin
biochemistry, protein isolation, purification and characterization, or
plant molecular biological techniques. For the first position,
experience in subcellular organelle isolation, purification and
characterization would be helpful
AVAILABLE: Oct 1995
SEND: CV and three letters of recommendation
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POSITION OFFERED: Post-Doc
CONTACT: Terry Bricker, Dept. of Microbiology, Louisiana State University,
Baton Rouge LA 70803, U.S.A. E-MAIL: Btbric@Lsuvm.Sncc.Lsu.Edu
RESEARCH: Structure and function relationships in photosynthesis
SEND: CV and three letters of recommendation
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POSITION OFFERED: Post-Doc
CONTACT: P. Sebban, Photosynthese Bacterienne, Bat. 24, Centre de Genetique
Moleculaire, CNRS, 91198, Gif FRANCE. TEL: 33-1-69-82-38-26
FAX: 33-1-69-82-35-62 E-MAIL: Sebban@Citi2.Fr
RESEARCH: Electrostatic effects and proton conduction in bacterial reaction
center membrane proteins.
REQUIREMENTS: Well-organized and flexible candidate able to pursue a
multidisciplinary approach. Desirable but not definitely needed is
experience in biochemistry and spectroscopy and knowledge of molecular
biology and genetics.
AVAILABLE: From 1 Oct 1995 for three years
SEND: CV and statement of research interests
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POSITION OFFERED: Post-Doc
CONTACT: David Kramer, Institute of Biological Chemistry, Washington State
University, Pullman WA U.S.A. TEL: 217-244-8913 or 217-333-7407,
E-MAIL: Kramer@Nemo.Life.Uiuc.Edu
RESEARCH: Characterization of photosynthetic electron transfer reactions in
intact higher plants and in evolutionarily interesting algal and
bacterial species.
REQUIREMENTS: U.S. citizenship or residence status. Experience desirable in
one or more of following: isolation of membrane protein complexes,
optical or electronics instrumentation, EPR spectroscopy, knowledge of
photosynthetic or respiratory electron transfer reactions.
AVAILABLE: 1 Sept 1995
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POSITION OFFERED: Post-Doc
CONTACT: H.Y. Yamamoto, University of Hawaii, 3050 Maile Way, Gilmore 202 B,
Honolulu, HI 96822 U.S.A. E-MAIL: Yamamoto@Uhunix.Uhcc.Hawaii.Edu
RESEARCH: Molecular biology and physiological function of violaxanthin de-
epoxidase, a key enzyme in the xanthophyll-dependent non-radiative
energy dissipation of excess energy to down-regulate PSII photochemical
efficiency.
REQUIREMENTS: self-motivated individual with a strong background in molecular
biology and publication record sought. Knowledge and interest in
photosynthesis highly desirable. Ph.D. in plant physiology,
biochemistry, molecular biology, or related discipline required.
SEND: CV and names of three references
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POSITIONS OFFERED: Two post-doc openings
CONTACT: Sabeeha Merchant, Department of Chemistry and Biochemistry, UCLA, 405
Hilgard Avenue, Los Angeles, CA 90095-1569. Tel: 310-825-8300,
Fax: 310-206-1035, E-mail: Merchant@Chem.Ucla.Edu
RESEARCH (Position 1): Copper-responsive gene expression in the context of
adaptation to copper-deficiency and the assembly of the photosynthetic
apparatus [EMBO J (1991) 10:1383; EMBO J (1995) 14:857; Plant Cell
7:623]
REQUIREMENTS (Position 1): Formal education and research experience in
biochemistry, molecular biology, or genetics.
RESEARCH (Position 2): Cytochrome biogenesis with an emphasis on the isolation
and functional analysis of genes involved in the specification of
cofactor (heme) -apoprotein assembly [EMBO J (1992) 11:2789; J Biol Chem
269:5824; Mol Gen Genet (1995) 246:156].
REQUIREMENTS (Position 2): Research experience in biochemistry, molecular
biology or genetics.
SEND: CV, publication list, relevant reprints, and letters of recommendation
****** POSITIONS SOUGHT ******
POSITION SOUGHT: Visiting professor/scientist (for 2-3 weeks only).
CONTACT: L.V. Venkataraman, "Sudarshana", #236, 8th Cross, Gokulam 3rd Stage,
Mysore 570 002 INDIA. TEL: 821-510006, FAX: 821-512539,
TELEX: 0846-320 POLY IN
RESEARCH EXPERIENCE: 25 years, basic applied aspects of Spirulina effluent
treatment, integrated systems, biotransformations, bioenergy production.
Over 200 publications.
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TRANSITIONS*TRANSITIONS*TRANSITIONS*TRANSITIONS*TRANSITIONS*TRANSITIONS*TRANSI
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MASAHIRO ISHIURA, TAKAO KONDO, and the rest of the circadian rhythm team
formerly of National Institute for Basic Biology in Okazaki has moved to
Nagoya University, where they will continue studying cyanobacterial clocks.
Department of Biology, Faculty of Science, Nagoya University, Furo-cho,
Chikusa-ku, Nagoya, 464-01 JAPAN. Tel: 81-52-789-2495, Fax: 81-789-2963,
E-mail: ishiura@Bio.Nagoya-U.Ac.Jp
L.V. VENKATARAMAN has taken an early retirement from the Central Food
Technological Research Institute in Mysore, India. He is keeping his academic
research alive, continuing to guide research students and consulting on
Spirulina biotechnology in India and abroad (see ANNOUNCEMENTS).
****** DAVID LAUDENBACH ******
We announce with great sadness that David Laudenbach died during surgery
on Thursday June 15, 1995 at the age of 35. It is always sad to lose a
colleague, and especially sad to lose a colleague who is so young and such an
integral part of the cyanobacterial community.
David received his MSc and PhD at the University of Toronto. His
research there focussed on the molecular genetic responses of Synechococcus
PCC 7942 to iron deficiency. He isolated the gene for flavodoxin and showed
that it was the second open reading frame of a dicistronic message whose
transcription was tightly regulated by iron. He demonstrated that the first
open reading frame encoded a protein with high homology to CP43, which he
correctly guessed to be the iron-stress-induced, PS II, chlorophyll-binding
protein that had been previously discovered in Lou Sherman's laboratory. He
also cloned the gene for ferredoxin and showed that its expression was not
affected by the concentration of iron in the growth medium. Before graduating
David isolated and created mutants for the genes encoding iron superoxide
dismutase and cytochrome c553. The productivity of his graduate years set a
pattern that would continue throughout the remainder of his career.
David left Toronto to do postdoctoral research at the Carnegie
Institution for Plant Science, Stanford University. His project concerned the
acclimation of Synechococcus to sulfur deficiency. David was able to
functionally define systems involved in sulfate transport and sequence the
genes encoding the components of these systems. He also defined a novel sulfur
limitation induced gene, designated rhd, that may be involved in the
utilization of certain thiol compounds during sulfur-limited growth. Finally,
Dave discovered the regulatory gene cysR and postulated its involvement in
controlling the utilization of thiocyanate during sulfur limitation. This work
was extended to some of the highly productive projects that Dave developed
independently as an Assistant Professor at the University of Western Ontario.
David was not the type of scientist that could be satisfied with one
project and his curiosity always got the better of him. For example, while at
Carnegie he started up collaborations with Dave Fork and Steve Herbert on the
acclimation of Synechococcus to oxidative stress and its affect on the
photosynthetic apparatus. His constant probing and 'playing' in the laboratory
provoked both new ideas and the development of new projects. David was a
talented and unique scientist.
David is survived by his wife Lori and two children Adam (5) and Theresa
(3). A fund has been established for Adam and Theresa. If you wish to
contribute please make cheques payable to "Lori Laudenbach in trust" and mail
them to CIBC, 228 Oxford Street East, London, Ontario N6A 1T7, Canada. Enclose
a letter stating who is making the contribution, including the names and
addresses of all for group contributions, and that the contribution is to be
directed to the trust fund for Adam and Theresa Laudenbach.
Arthur Grossman & Neil Straus
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NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEWS*NEW
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ALLEN ACCLAIMED BY NATIONAL SOCIETY
Mary Mennes Allen was honored at the 1995 meeting of the American
Society for Microbiology with the Carski Foundation Distinguished Teaching
Award, in recognition of her career in inspiring undergraduates towards a
career in science. Needless to say, a useful tool in her inspirational efforts
has been her ongoing research into the function of cyanophycin in
cyanobacteria.
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CONTROLLED EXPRESSION OF FOREIGN GENES IN CHROMOSOMES OF SYNECHOCOCCUS
A few years ago workers at the University of Utrecht described a novel
method to facilitate the stable insertion of foreign genes into an innocuous
locus of the chromosome. The constructed chromosomal location, called PIM (for
platform for integration in metF), consists of a promoterless bla gene and
oriV, both from pBR322, a plasmid commonly used in molecular biology.
Insertion of genes of interest into that site could be achieved, with
selection for ampicillin (encoded by bla), upon recombination between the
platform and their pBR322-derived vector. Dirk Geerts now tells us that he and
others in Utrecht have extended the technique to permit inducible, high-level
expression of genes placed in the platform. The original method has further
been modified to greatly reduce the aberrant recombinational events that had
plagued the technique in the past.
Their new vector, pTrcIS, provides a strong trc promoter whose
expression is well controlled by the lactose repressor, encoded by the lacIq
gene also on the plasmid. Downstream from this promoter is the lac operon
ribosome binding site and a polylinker to facilitate transcriptional or
translational fusions of inserted genes. Of particular utility is an NcoI site
for the insertion of the 5'end of a gene directly to the ATG start codon.
Flanking this region are a complete version of bla and oriV, required for
integration into the platform. The Utrecht group also place aadA, determining
resistance to streptomycin and spectinomycin.
They found, using petE (encoding the precursor to plastocyanin) from
Anabaena PCC 7937 (Anabaena variabilis), and uidA (encoding beta-
glucuronidase, GUS) from E. coli, that double recombination events placing the
foreign gene into the platform occurred with a very low incidence of false
positives when streptomycin was used as the selective agent. When ampicillin
alone was used, the number of colonies recovered was much higher but the
majority of recombinants were not true double recombinants, and the frequency
with which the foreign gene was expressed in the recombinant varied from 0 to
100%, depending on the insert. Selection for streptomycin evidently ensures
that virtually all of the colonies resulting from transformation of
Synechococcus have the desired phenotype.
Expression of the foreign gene could be controlled within a wide dynamic
range by the addition of graded amounts of the lac inducer IPTG, with full
repression in the absence of the inducer. The highest level of induction was
36-fold, as judged by expression of GUS, or 100-fold, as judged by expression
of petE. The level of expression by the Ptrc-uidA fusion is almost 4-fold
higher than that achieved by the strongest the cyanobacterial promoter (PpetF)
tested.
The work has recently been fully described [Geerts et al (1995)
Microbiol-UK 141:831-841].
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EXCRETION OF AMMONIA FROM IMMOBILIZED ANABAENA EXPLAINED
Symbiotic cyanobacteria commonly release fixed nitrogen resulting from
N2 fixation to their hosts. It has long been a hope of some that cyanobacteria
could be induced to excrete ammonia on a large-scale industrial setting. In
1987, Shi Ding-Ji and others [Planta 172:298-308] reported that ammonia
excretion occurred simply by N2-fixing cyanobacteria immobilized within
polyurethane foams. Why this should be the case has been a mystery, but Shi
describes to us how Duan Xue-Yan and others at Capital Normal University and
Academia Sinica in Beijing have brought us one step closer to its solution.
The Beijing group found that Anabaena sp. strain 2B (isolated as an
epiphyte of Azolla caroliniana) immobilized within polyurethane showed higher
(1- to 2-fold) glutamine synthetase (GS) activity than a free-living culture
over the several days following initiation of the experiment. Over the longer
term, however, GS activity in immobilized Anabaena drops 10-20% below that of
the free-living culture. This period of low GS activity roughly corresponds
to the period of high production of ammonia by immobilized Anabaena reported
earlier.
Hybridization of mRNA isolated from free and long immobilized cultures
to a probe specific for GS mRNA indicate that the drop in GS activity is due
to a corresponding decrease in GS message. In order to achieve this result,
the group had to improve upon existing methods to extract RNA from immobilized
cyanobacteria. Their modification permits efficient isolation, as judged by
comparison of stable rRNA from immobilized and free cultures.
Details of the work have been published [Duan et al (1994) Chinese J Bot
6:102-106 (English)].
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X INTERNATIONAL CONGRESS ON NITROGEN FIXATION - MEETING REPORT
The 10th International Congress on Nitrogen Fixation took place 28 May
to 3 June of this year in St. Petersburg, Russia. While the majority of
presentations concerned themselves with the doings of heterotrophic bacteria,
there were a few cyanobacterial nuggets, some of which are reported below.
Bernd Masepohl (Bonn) reported the identification of a NOVEL REPEATED
DNA ELEMENT in Anabaena PCC 7120 with so far unknown function. This long
tandemly repeated repetitive (LTRR) sequence is 37 bp long and contains an
inverted repeat sequence. An LTRR-specific probe hybridized to numerous DNA
regions in Anabaena PCC 7120 and many other cyanobacteria.
In addition he described the construction of a mutant derivative of
Anabaena PCC 7120 defective in the FERREDOXIN-encoding gene, fdxH. The mutant
exhibited much reduced nitrogenase activity, confirming that this [2Fe-2S]
heterocyst ferredoxin (see B. Schrautemeier, below) is the principal electron
donor to nitrogenase, but may also partly be replaced by (an) alternate
donor(s).
The mechanism of cyanobacterial NITROGENASE REGULATION OR MODIFICATION
into the inactive form of the Fe-protein is still an enigma, according to John
Gallon (Swansea). ADP-ribosylation is evidently not involved, in contrast to
the importance of such a modification in the case of glutamine synthase, as
recently reported by Noel Carr and Nick Mann.
The mechanism of OXYGEN SENSING is now better understood in Azotobacter
vinelandii (if we may slip in a noncyanobacterium). Ray Dixon (Sussex)
described the characterization of the oxygen-sensor protein NifL. It contains
FAD with FMN as minor component and controls the catalytic activity of NifA
in the active ADP-bound form.
The regulatory protein OxyR plays a role under OXIDATIVE STRESS in
E. coli and Salmonella typhimurium. Karin Jaeger (Hannover) found an OxyR-like
protein in Anabaena variabilis and Anabaena PCC 7120 by using a specific
antibody against the E. coli protein. Southern blot analyses with the E.coli
gene probe reveled no signal with cyanobacterial genomic DNAs.
Bernhard Schrautemeier (Bonn) reported on DUAL MO-NIF SYSTEMS expressed
from separate nif gene clusters (nif1, nif2) of Anabaena variabilis ATCC 29413
that also were independently discovered by Teresa Thiel and coworkers in St.
Louis. Teresa, using a lacZ reporter system with a fluorescent substrate,
conclusively demonstrated localized expression of nif1 (only in heterocysts)
versus nif2 (in all vegetative cells), Bernhard's approach emphasizes the time
component/differential kinetics of oxygen-controlled nif2- versus
developmentally controlled nif1-expression after nitrogen deprivation: Nif2
is expressed only under strictly anaerobic conditions as early as 1-2 hours
after nitrogen stepdown -- long before the appearance of proheterocysts. In
contrast, Nif1 is expressed only after (pro)heterocysts have appeared, i. e.
not earlier than about 10 hours after nitrogen depletion, irrespective of
anaerobic or aerobic growth conditions. By using a standardized comparative
induction assay monitoring nitrogenase activity during the 20 hours following
nitrogen deprivation, he additionally demonstrated that Anabaena PCC 7120 has
no characteristics of a functional Nif2 system.
Examining the region upstream from each nifHDK cluster, Bernhard hit
upon different genes encoding ferredoxins: fdxH1 and fdxH2 for the nif1 and
nif2 clusters, respectively. It is interesting that FdxH1 is oxygen-tolerant
in vitro, while FdxH2 is rapidly inactivated by oxygen. Both are equally
effective in donating electrons to nitrogenase isolated from heterocysts. The
fdxH2 gene, but not fdxH1, is followed by a gene, fdxB, encoding a second
ferredoxin of unknown function, as also present downstream of fdxH from the
nonheterocystous filamentous Plectonema PCC 73110. Hence the Nif2 system is
homologous to the single, environmentally regulated Mo-Nif system expressed
in all cells of nonheterocystous filamentous species [Smoker et al (1990) Meth
Molec Cell Biol 2:59-65].
Many additional questions now arise from the work of Bernhard, Teresa,
and coworkers. In particular: (a) How do nif1 and nif2 differ in their
regulatory mechanisms yet intersect in their dependence on nitrogen
deprivation? (b) What is the distribution of the two systems amongst nitrogen-
fixing cyanobacteria? (c) What is the benefit of two coexisting nif systems?
-- Karin Jaeger
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EUROPEAN WORKSHOP ON THE MOLECULAR BIOLOGY OF CYANOBACTERIA - MEETING REPORT
****** BIOENERGETICS AND PHYSIOLOGY ******
Several presentations related to the structure and function of
FERREDOXIN-DEPENDENT ENZYMES. Data from Herbert Boehme's group in Bonn
suggests a common but not identical ferredoxin binding domain on the
ferredoxin-dependent enzymes nitrate reductase, nitrite reductase and
ferredoxin NADP-reductase (FNR). It is noteworthy that the same mutations in
specific amino acids of ferredoxin that stimulated the reversed flow from
NADPH-reduced FNR to oxidized ferredoxin also severely inhibited electron
transport from reduced ferredoxin to FNR. Carlos Gomez-Moreno (Zaragoza)
presented rapid kinetic characterizations of ferredoxin, flavodoxin and FNR
mutants in which amino acids involved in the interaction between these
proteins had been modified. Two others from Zaragoza, Maria Fillat and Marisa
Peleato, described, respectively: (1) the overexpression in a
protease-deficient E. coli of the 49-kDa form of FNR (the complete product of
petH) from Anabaena and (2) the characterization of different
FNR-phycobiliprotein complexes of Anabaena, isolated from vegetative cells and
heterocysts.
CAROTENOID BIOSYNTHESIS was the focus of presentations by Gerhard
Sandmann (Frankfurt) and Blanca Fernandez (Sevilla). Sandmann reported the
cloning of the zeta-carotene desaturase gene from Anabaena PCC 7120 by
heterologous complementation. Fernandez, using the cat gene as a reporter,
showed stimulation of the expression of the phytoene desaturase (crtP)
promoter at high light intensities, a result in agreement with the
photoprotective role ascribed to carotenes.
The molecular bases of the ADAPTIVE RESPONSES of cyanobacteria to
changes in light conditions were addressed by a few presentations. Jean
Houmard (Paris) reported that whereas changes in the photosynthetic photon
flux density exerts a major influence on the differential expression of genes
within the psbA and psbD multigene families, changes in light wavelength also
result in profound modifications of the light harvesting apparatus, in those
strains that exhibit chromatic adaptation. In Calothrix PCC 7601, different
phosphorylated DNA-binding proteins, namely RcaA and B (which specifically
bind to the promoter region of the phycoerythrin operon) and RcaD (which binds
to the phycocyanin-2 operon), are expressed under green-light and red-light,
respectively. However, no difference was found in the subunit composition of
the RNA polymerase isolated from cells grown under the two conditions.
Insights into the LIGHT REGULATION of the PSBA GENE FAMILY (encoding D1
protein) in Synechocystis PCC 6803 were provided by Christer Jansson
(Stockholm) and David Campbell (Umea). Jansson found that psbA2 and psbA3, two
gene copies differing in their promoter elements, were light-regulated and
transcribed at vastly different levels (30-fold higher for psbA2).
Inactivation of psbA2 by in vitro mutagenesis led to an 8-fold up regulation
of psbA3 gene transcription. Site-specific mutagenesis permitted the
identification of putative Mn-binding amino acids within D1 and sequences
involved in the light-triggered proteolytic degradation of this protein.
Campbell reported that Synechococcus responds to excitation stress by
replacing the constitutive form of the D1 protein (D1:1) by another form,
D1:2, that confers increased resistance to photoinhibitory damage and a higher
photochemical efficiency of PS II. This D1 exchange is a response to excess
excitation of photosynthetic electron transport, and not a specific response
to light intensity per se.
Aaron Kaplan (Jerusalem) described novel HIGH-CO2-REQUIRING MUTANTS of
Synechococcus PCC 7942. These mutants were obtained upon integration of
plasmids containing DNA internal to the CO2-related operons, selected from a
library composed by short genomic fragments. The mutant-forming plasmids were
retrieved and their genomic regions used as probes of wild-type transcripts
and genomic DNA. By this means, Kaplan's group showed that the different
regions are transcribed and do not lie in close proximity to each other in the
chromosome. The study by fusion experiments of the promoter regions of genes
modulated by changing CO2 concentration indicated the presence of enhancing
and repressing elements. Conserved sequences were found in the promoter region
of several CO2-responding genes. Francoise Joset's group (Marseille) described
the characterization of a gene from Synechocystis PCC 6803, hatR, involved in
a high affinity system of HCO3- uptake and the identification of proteins
showing different levels of synthesis in response to changes in the levels of
inorganic carbon.
Aurelio Serrano (Sevilla) cloned the NAD(P)-dependent GLYCERALDEHYDE
3-PHOSPHATE DEHYDROGENASE (G3P DHase) from Synechocystis PCC 6803, by
complementation of an E. coli gap- mutant with a genomic library. The enzyme
restored the glycolytic pathway in E.coli, and thus may be presumed able to
function in that capacity in Synechocystis as well. Since G3P DHase is already
known to be essential in the reductive pentose phosphate pathway, the enzyme
may therefore play both anabolic and catabolic roles in Synechocystis. The
sequence of the complementing gene predicted a protein very similar (70-80%
identity) to G3P DHase from chloroplasts of higher plants. Southern blots
using as probes the cloned G3P DHase genes of Synechocystis and E. coli
indicated that two genes, one corresponding to each type, are present in the
Synechocystis genome. However, since immunological and biochemical data are
consistent with the presence in Synechocystis, of only an NAD(P)-dependent
enzyme, Serrano suggested that the E. coli-like gene may be a pseudogene or
a gene not expressed under normal culture conditions.
Two presentations had important implications regarding cyanobacterial
RESPIRATION. Georg Schmetterer (of Vienna) obtained cox- mutants of
Synechocystis PCC 6803 in which the three genes coding for the terminal
oxidase of aa3 type were inactivated. Surprisingly, although no cytochrome c
oxidation by membranes of the mutants was observed, the intact cells respire
almost normally. Schmetterer explained these results by postulating the
existence of An "alternative terminal oxidase", sensitive to KCN, that reduces
O2 in the dark with NAD(P)H. Gunther Peschek (Vienna) presented results
indicating that the cyanobacterial cytochrome c oxidase might be subject to
adenylate regulation. A putative mitochondria-like subunit IV gene (ctaIV) was
identified in Synechocystis exhibiting consensus sequences of
adenylate-binding enzymes.
Norio Murata (Okazaki) described very interesting results on the GENETIC
MANIPULATION OF MEMBRANE LIPIDS in cyanobacteria. His group was able both to
decrease the degree of unsaturation of fatty acids in Synechocystis PCC 6803
(by inactivating the corresponding desaturase genes) and to increase the
degree of unsaturation of fatty acids in membranes of Synechococcus PCC 7942
(by transformation with foreign cyanobacterial desaturase genes). These
changes did not affect rates of photosynthesis and photosynthetic electron
transport and only scarcely affected heat stability of oxygen evolution.
However, a lower degree of unsaturation enhanced photoinhibition at low
temperatures and a higher degree of unsaturation accelerated recovery from
photoinhibition. These results may help to elucidate the mechanisms of
photoprotection of photosynthetic organisms at low temperatures.
-- Aurelio Serrano
****** NITROGEN REGULATION/METABOLISM AND N2 FIXATION ******
Antonia Herrero and Ignacio Luque (both of Sevilla) reported results
concerning the mechanism of GENE REPRESSION BY AMMONIA in Synechococcus
PCC 7942. They found that mutant strains that express NtcA to a high and
constitutive level still require the absence of ammonium to express
NtcA-regulated genes (e.g. nir operon, glnA). They suggested that a
coactivator may be required for the expression of nitrogen-regulated genes or
that NtcA protein is posttranscriptionally interconverted between an active
and an inactive form in response to the nitrogen status. Antonia also proposed
that the level of NtcA might contribute to the differential regulation of some
genes through weak binding of the protein to sites deviating from the known
NtcA consensus binding site.
Jose Frias (Sevilla) discussed the GENES OF NITRATE assimilation: their
regulation and function. The nir-nrtABCD-narB gene cluster (the nir operon)
of Anabaena PCC 7120 was cloned and analyzed. Northern analysis showed that
the nir operon is transcribed in the absence of ammonium with or without
nitrate or nitrite in the medium, this despite the fact that high levels of
nitrate and nitrite reductase activities occur only in the former case. A 460
bp leader sequence between the Ntc-regulated promoter and the first codon of
nir seems to lack any function: when removed no change in phenotype was
observed. Insertional inactivation of nrtA resulted in mutants that were
unable to transport nitrate at low external concentration (0.1 mM), but at
high concentration (18 mM) nitrate was taken up at a slow rate and reduced
to ammonium. High activities of the nitrate assimilation enzymes were observed
at either level of nitrate concentrations but neither could repress heterocyst
formation.
Paco Navarro (Sevilla) found two different genes, gltS and gltB, in
Synechocystis PCC 6803, that encode ferredoxin-dependent GLUTAMATE SYNTHASES
(GOGAT). Inactivation of either one did not significantly impair growth
(concomitant inactivation of both has not yet been tried). While gltS was
present in many other cyanobacteria tested, gltB was additionally found only
in Pseudoanabaena PCC 6903. Both enzymes expressed in E. coli accept electrons
from PetF-type ferredoxins, but flavodoxin was inactive. Interestingly, GltB
was equally active with heterocyst ferredoxin (FdxH). Figueroa (Sevilla)
cloned gltS from Anabaena PCC 7120. GltS activity was highest in crude
extracts of cells using N2 as the nitrogen source, as opposed to nitrate or
ammonium, hinting at a role for this GOGAT in heterocyst metabolism.
Reyes and Florencio (Sevilla) reported that the REDOX STATE controls the
transcription of glnA, encoding GLUTAMINE SYNTHETASE (GS). Transcript
abundance was high when Synechocystis PCC 6803 was grown in the light or in
the dark with glucose and low in the dark without glucose or when DCMU (a
PSII-inhibitor) or DBMIB (a cytochrome b6/f complex-inhibitor) was added.
N-starvation provoked a delay in decrease of glnA transcripts suggesting a
connection between nitrogen and redox controls of transcript levels. Crespo
(Sevilla) found that redox control seems also to govern inactivation of
glutamine synthetase (GSI) in vivo. In this case, however, the addition of
DBMIB, leading to a reduced plastoquinone (PQ) pool, was not inhibitory. This
result suggests that the redox state of PQ or a component of the b6/f-complex
is a signal for modification.
The same group also characterized a SECOND TYPE OF GLUTAMINE SYNTHETASE
from Synechocystis PCC 6803, encoded by glnN, similar to the GSIII-type
enzymes found in the Bacteriodaceae. GlnN has a larger subunit size (75kD)
than the 50kD product of glnA and, unlike the dodecameric GlnA, probably
exists in its native state as a hexamer. According to Western blot analysis,
GSIII is more abundant in PCC 6803 and other non nitrogen-fixing cyanobacteria
when they are starved for nitrogen. GSIII is lacking, however, in N2-fixing
Anabaena PCC 7120.
Nicole Tandeau de Marsac and coworkers (Paris) described PII PROTEIN as
the central node for the coordination of nitrogen and carbon assimilation in
cyanobacteria. PII is a protein whose homologue in enteric bacteria is
involved in regulation of GS activity and Ntr-regulated gene expression. She
reported that a PII-deficient mutant of Synechococcus PCC 7942 can take up
nitrate even in the normally inhibitory presence of ammonium. The mutant has
also lost the ability to adapt rapidly to changes in light, nitrogen, and
carbon supplies. PII thus functions to integrate nutritional stimuli and to
reestablish a proper C/N-ratio for balanced cell growth. In Calothrix
PCC 7504, PII is found to be unmodified during the hormogonial stage of
growth, whereas PII modification is most pronounced under conditions of
heterocyst differentiation. Thus, in filamentous strains PII may be
additionally involved in cell differentiation processes.
Karl Forchhammer (Munich) devised an in vitro test for PHOSPHORYLATION
OF PII that clearly demonstrated that 2-ketoglutarate is sufficient to
activate PII kinase from Synechococcus PCC 7942. No other compound tested
(e.g., glutamine or other amino acids) could substitute or counteract the
stimulation by 2-ketoglutarate. The latter may serve as an intracellular
signal to monitor the balance of assimilated carbon and nitrogen that is
sensed by PII kinase and transmitted to PII by protein serine phosphorylation.
Lucas Stal (Amsterdam) made an interesting observation related to the
CAPABILITY OF NONHETEROCYSTOUS CYANOBACTERIA TO FIX NITROGEN predominantly in
the light. He noted that a Cyanothece strain is impaired in nitrogen fixation
when grown in batch cultures, where they produce sulfated extracellular
polysaccharides and thus rapidly deplete the medium of sulfate. In continuous
cultures with a continuous supply of sulfate nitrogenase activity was high and
confined predominantly to the light. The same was true when sulfate was added
to a sulfate-depleted batch culture. This intriguing observation leaves us
once more perplexed (as with the case of Trichodesmium): how do they do it
without heterocysts?
-- Bernhard Schrautemeier
****** ECOLOGY ******
One perennial problem for ecologists is that of IDENTIFYING the
ORGANISMS present in natural populations. Strain identification is also a
problem for those of us working on newly isolated strains. A variety of
molecular techniques are available for strain identification and
discrimination; the results presented on three posters (Anneliese Ernst,
Konstanz; Gary Barker, Bristol; Suzzanne McColl, Liverpool) lend yet more
evidence to what has been long suspected, namely that natural populations of
cyanobacteria consist of many distinct clones.
The biology of GAS VACUOLATE CYANOBACTERIA was covered in two
presentations. The advantages accruing from gas vesicle production by
Aphanizomenon in the Baltic Sea is being quantified by Walsby and co-workers
(Bristol); such colonial forms can gain a 3-fold photosynthetic advantage over
their non-buoyant competitors by rapidly moving back towards surface, and
light, after mixing-events. The genes involved in producing gas vesicles and
the interactions between the gene products were described by Hayes et al. Once
thought to be a simple structure formed by self assembly of a single type of
protein, it is now clear that it takes the concerted action of at least six
different gene products to assemble these structures.
Elke Dittmann et al. (Berlin) described progress toward the complete
characterization of the genes encoding PEPTIDE SYNTHETASES of cyanobacteria.
These incredibly complex enzymes are responsible for the synthesis of the
cyclic peptide toxins. The group in Berlin have partially characterized a gene
from Microcystis aeruginosa using conserved domains from peptide synthetases
to provide probes. This approach is similar to that described by Leo
Rouhiainen et al. (Helsinki) in Urbino for the genes from Nodularia. With the
genes available from a number of organisms it should now be possible to study
the biological role of these toxic compounds.
Molecular mechanisms of SALT TOLERANCE were described in two
presentations (Martin Hagemann and Ellen Zuther, Rostok). A total of 18 salt
sensitive mutants of Synechocystis PCC 6803 were produced by random cartridge
mutagenesis; 9 of these were unable to synthesize glucosylglycerol. One of the
genes identified, stpA, had been previously characterized (Francoise Joset,
Marseille). Three other ORFs have been characterized in Rostock; the role of
theses genes has yet to be confirmed but glucosylglycerol transport and
positive regulation of glucosylglycerol synthesis seem likely candidates.
Nigel Robinson (Newcastle) gave a lucid summary of work carried out
in his laboratory on the regulation of expression of the METALLOTHIONINE-
ENCODING GENE smtA from Synechococcus PCC 7942. SmtB is a repressor of smtA
expression that dissociates from the smtA promoter in the presence of Zn2+.
Upstream of smtB is smtZ, a gene that encodes a protein the C-terminal end of
which has the features of a zinc-finger: SmtZ may up-regulate smtA expression
in the presence of Zn2+. Upstream again is dnaG, encoding primase which could
be a zinc metalloprotein. An octameric palindrome HIP1 (5'- GCGATCGC-3') is
involved in the deletion of smtB in cells selected for zinc tolerance. This
sequence occurs at much higher than expected frequencies in many cyanobacteria
(but not in any of the marine isolates investigated); is it involved in genome
plasticity? We will have to wait for the answer to that question.
All of the presentations at the meeting were excellent and I wish I
had the time to write about them all (in particular Nick Mann, Warwick, gave
a first class talk of extreme ecological relevance, but mine came straight
afterwards so I missed most of what he saying) but at least you now have the
gist of some of the topics covered.
- Paul Hayes
==============================================================================
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==============================================================================
****** EVOLUTION and ECOLOGY ******
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****** SYMBIOSIS ******
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****** TOXINS and NATURAL SUBSTANCES ******
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****** TOXINS and NATURAL SUBSTANCES (Physiological Effects) ******
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Papadogiannakis N (1995). (-)-Indolactam V-induced mitogenesis in human fetal
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****** PHYSIOLOGY ******
Bonilla I, Bolanos L, Mateo P (1995). Interaction of boron and calcium in the
cyanobacteria Anabaena and Synechococcus. Physiol Plant 94:31-36
Evans DJ, Evans DG, Lampert HC, Nakano H (1995). Identification of four new
prokaryotic bacterioferritins, from Helicobacter pylori, Anabaena
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Muniz WH, Stevens SE (1994). Development of motility in cultures of the
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of a Synechococcus sp. strain PCC 7942 polypeptide structurally similar
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Sudo H, Burgess JG, Takemasa H, Nakamura N, Matsunaga T (1995). Sulfated
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Waterbury J, Ostroumov SA (1994). Effect of non-ionogenic surfactant on
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****** MEMBRANES & LIPIDS ******
Avelange-Macherel MH, Macherel D, Wada H, Murata N (1995). Site-directed
mutagenesis of histidine residues in the þ12 acyl-lipid desaturase of
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Kanervo E, Aro EM, Murata N (1995). Low unsaturation level of thylakoid
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Malakhov MP, Los DA, Wada H, Semenenko VE, Murata N (1995). Characterization
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8
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****** STRESS RESPONSES ******
Buck DP, Smith GD (1995). Evidence for a Na+/H+ electrogenic antiporter in an
alkaliphilic cyanobacterium Synechocystis. FEMS Microbiol Lett 128:315-
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Tadros MG, Smith W, Joseph B (1995). Yield and analysis of cyanobacteria
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****** NITROGEN METABOLISM ******
Cohen-Kupiec R, Zilberstein A, Gurevitz M (1995). Characterization of cis
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Forchhammer K, Demarsac NT (1995). Functional analysis of the phosphoprotein
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Jahns T, Schafer U, Kaltwasser H (1995). Heat-stable ureases from two
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Kashyap AK, Shaheen N, Prasad P (1995). Characteristics and regulation of the
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Singh S, Bisen PS (1994). Assimilation of ethylenediamine as nitrogen source
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Suzuki I, Horie N, Sugiyama T, Omata T (1995). Identification and
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Plant Physiol 107:791-796
****** NITROGENASE and DIFFERENTIATION ******
Lyons EM, Thiel T (1995). Characterization of nifB, nifS, and nifU genes in
the cyanobacterium Anabaena variabilis: NifB is required for the
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Bauer CC, Buikema WJ, Black K, Haselkorn R (1995). A short-filament mutant of
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Bauer CC, Haselkorn R (1995). Vectors for determining the differential
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****** CARBON METABOLISM ******
Jensen TE (1994). Fine structure of elongate polyhedral bodies (carboxysomes)
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Moezelaar R, Demattos MJT, Stal LJ (1995). Lactate dehydrogenase in the
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Summers ML, Meeks JC, Chu S, Wolf RE (1995). Nucleotide sequence of an operon
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****** PHOTOSYNTHESIS and PHOTOSYSTEMS ******
Govindjee (1995). Sixty-three years since Kautsky: Chlorophyll a fluorescence.
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Hovenden MJ, Seppelt RD (1995). Utility of modulated fluorescence in measuring
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****** PHYCOBILISOMES and OTHER PIGMENTS ******
Bastia AK, Adhikary SP (1995). A phycoerythrin-lacking mutant induced by DCMU
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****** ELECTRON TRANSPORT and BIOENERGETICS ******
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Varley JPA, Moehrle JJ, Manasse RS, Bendall DS, Howe CJ (1995).
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****** MOLECULAR GENETICS, EPISOMES, AND METABOLISM OF MACROMOLECULES ******
Churin YN, Shalak IN, Borner T, Shestakov SV (1995). Physical and genetic map
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Zaug AJ, Davilaaponte JA, Cech TR (1994). Catalysis of RNA cleavage by a
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Nagaraja R, Haselkorn R (1994). Protein HU from the cyanobacterium Anabaena.
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****** APPLIED CYANOBACTERIOLOGY ******
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Growth of Phormidium sp in aerobic secondary piggery waste-water. Appl
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J Appl Bacteriol 78:194-199
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AUSTRALIA Steve Delaney Department of Biotechnology,
/NEW ZEALAND University of New South Wales, P.O.
Box 1, Kensington, New South Wales
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INDIA Joe Thomas Biotechnology Division, SPIC Science
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ISRAEL Elisha Tel-Or Dept. of Agricultural Botany, The
Hebrew University, Rehovot 76100
(Tel) 08-481262
ITALY Mario Tredici Departimento di Scienze e Tecnologie
Alimentari e Microbiologiche.
Universita degli Studi di Firenze,
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Research, P.O.box 69 Korsvall, N-0808
Oslo 8 NORWAY. (Tel) 47 22 185266
(Fax) 47 22 185200
U.K. Tony Walsby Dept. of Botany, University of
Bristol, Bristol BS8 1UG.
(Tel) 0272-303030
ANYWHERE ELSE Jeff Elhai Dept. of Biological Sciences, Florida
International University, University
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(Tel) 305-348-3584, (Fax)305-348-1986
(E-mail) Cyano@Servax.Fiu.Edu