Bariloche protein symposium argentine society for biochemistry and molecular biology

70
BIOCELL, 27 (Suppl. I), 2003
MI-P42.
SUCROSE ENZYME EXPRESSION IN AN ANABAENA
MUTANT IMPAIRED IN NITROGEN FIXATION
Marcozzi, Clarisa; Cumino, Andrea C.; Curatti, Leonardo and
Salerno, Graciela L.
Centro de Investigaciones Biológicas (FIBA), Mar del Plata,
Argentina. E-mail: [e-mail]
Cyanobacteria constitute a diverse group of prokaryotes
characterized by their ability to perform oxygenic photosynthesis.
Most of them are able to use nitrate or ammonium as a nitrogen
source and some strains are capable of N
2 
fixation. Nitrogen control
is mediated by a unique transcription factor that acts as a global
regulator of nitrogen and carbon assimilation. Sucrose is involved
in the diazotrophic metabolism of the heterocyst-forming
cyanobacterium  Anabaena sp. PCC 7120. It is synthesized by
sucrose-phosphate synthase and sucrose-phosphate phosphatase,
and cleaved by sucrose synthase, enzymes encoded by spsA and
spsB,  spp, and susA respectively. During N
2
 fixation, sucrose
synthesis is increased and sucrose cleavage diminishes. The aim
of this work is to study the expression of the enzymes involved in
sucrose metabolism in Anabaena sp. PCC 7120 and in a mutant
strain (CSE2) lacking the global nitrogen transcriptional regulator
and the capacity of N
2 
fixation. Sucrose level was about 5-fold
lower in the mutant than in the PCC 7120 strain, 24 h after
combined-nitrogen step-down. Accordingly, expression in the CSE2
strain of spsB decreased, while of susA increased, both at the
mRNA and enzyme activity level. Moreover, there were detected
putative binding sites for the transcriptional regulator in the
promoter sequences of sucrose metabolism genes. Taken together
these results suggest that sucrose metabolism may be coordinated
with nitrogen assimilation through the action of the nitrogen
transcriptional global regulator.
Supported by Fundación Antorchas, Conicet, Univ. Nac. de Mar
del Plata and Fiba.
MI-P43.
VP7 AND VP4 GENOTYPING OF  BOVINE GROUP A
ROTAVIRUS
Masini Matilde
1
, Argüelles Marcelo
1
, Mattion Nora, Blackhall
Jorge
2
 and Glikmann Graciela
1
.
1
Univ. Nac. de Quilmes, Bernal, Bs As. 
2
Biogenesis S.A., Garin,
Bs. As. E-mail: [e-mail]
Group A rotaviruses, members of the Reoviridae family, are
important viral diarrheal agents and have been recognized as the
major etiologic agents of acute gastroenteritis in cattle (1969) and
young children worldwide (1973). Two outer capsid proteins, VP4
and VP7, elicit independently, neutralizing antibodies and specify
the virus P and G serotypes, respectively. At least, 14 G and 20 P
serotypes or genotypes of group A rotaviruses have been described
in humans and animals, being G6, G10, P1, P5 and P11 the most
common strains found in cattle.  Determination of the serotype
specificity and characterization of the genetic and antigenic
diversities of circulating bovine rotavirus strains (brv) in the field
is important to develop more efficacious vaccines. The genetic
variation among G types for human and bovine rotavirus strains
has been described, but little information is available for P types.
Group A rotavirus was unequivocally demonstrated in 95% of the
samples tested by enzyme-linked immunosorbent assay (ELISA)
for detection of VP6 protein, and reverse transcription-PCR (RT-
PCR) for amplification of the VP7 and VP4 genes. G and P typing
was carried out by nested amplification of variable sequences of
both genes using two G- and three P-type-specific primers. Results
obtained by this method showed the prevalence of the following
combinations: P[5]G6: 65%; P[11]G6: 14%; while mixed infections
with more than one type were found in 15% of the samples.
MI-P44.
AUTOANTIBODIES INDUCED IN MICE INFECTED WITH
MOUSE HEPATITIS VIRUS
Patricia A. Mathieu
1
, Karina A. Gómez
1
, Jean-Paul Coutelier
2
and Lilia A. Retegui
1
.
1
Facultad de Farmacia y Bioquímica, (UBA) and 
2
Christian de
Duve Institute of Cellular Pathology, Brussels, Belgium. E-mail:
[e-mail]
Mouse hepatitis virus (MHV) diversely affects immune response,
depending on the viral strain and the mouse genetic background.
We have shown that sera from mice infected with MHV A59
contained autoantibodies (autoAb) directed toward a 40 kDa
protein present in mouse liver and kidney extracts. Reactive
immunoglobulins were detected from 10 days up to 12 weeks after
infection. No correlation was found between the development of
hypergammaglobulinemia that followed viral infection and the
occurrence of the autoAb. The 40 kDa protein was purified from
mouse liver and identified as fumarylacetoacetate hydrolase (FAH).
To further characterise the autoimmune response, homologous
aminoacid sequences from FAH and viral proteins were synthesized
by the PEPSCAN technique, and each peptide was incubated with
individual sera from either MHV-infected or FAH-immunized
mice. Results indicated that three homologous sequences were
recognised by the autoAb induced by the viral infection, whereas
the anti-FAH samples reacted heterogeneously towards the various
peptides.  Moreover, ELISA and Western-blot competition
experiments indicated that the reactivity of sera from infected mice
depended on the conformational states of both FAH and MHV
proteins. These observations suggest that the autoAb elicited by
MHV-infection recognised mainly cryptic FAH epitopes and that
the response differed from one animal to other.
MI-P45.
ORNITHINE AND ARGININE  DECARBOXYLASE
ACTIVITIES AND EFFECT OF SOME POLYAMINE
SYNTHESIS INHIBITORS ON Gigaspora rosea
GERMINATING SPORES
Sannazzaro Analía
1
, Álvarez Cora
1
, Menéndez Ana
2,1*
, Pieckenstain
Fernando
1
, Albertó Edgardo
1
, Ruiz Oscar
1
.
1
Instituto Tecnológico de Chascomus (IIB-INTECH), CONICET-
Universidad Nacional de General San Martín. 
2
Departamento de
Biodiversidad y Biología Experimental, Facultad de Ciencias
Exactas y Naturales, Universidad de Buenos Aires.*E-mail:
[e-mail]
The aim of the present study was to increase the knowledge on
polyamine metabolism in AM fungi and to determine whether
inhibitors of polyamine synthesis (which could be used as
fungicides) may cause negative effects on spore germination and
hyphal growth of Gigaspora rosea.  In vivo decarboxylation of
radioactive substrates, showed that the ODC pathway is active
and ADC activity occurs in G. rosea spores. Results showed that
urease is active during spore germination and provided evidence
on the activity of NCPase, downstream the ADC. Effect of
polyamine synthesis inhibitors on free polyamine content of spores,
germination and germinating tube growth was studied. Spermine
and spermidine were reduced by DFMA at concentrations 0.05
and 0.5 mM, whereas CHA or DFMO did not affect spermine and
spermidine levels. DFMA and DFMO, either alone or in
combination, and putrescine, exerted no significant effects on spore
germination at any of the assayed concentrations. In contrast, CHA
reduced spore germination by far than 70%. On the base of our
results, the use of polyamine biosynthesis inhibitors to control
fungal pathogens would not affect germination and growth of G.
rosea.

---