3 research outputs found
Changes in Cathepsin Gene Expression and Relative Enzymatic Activity During Gilthead Sea Bream Oogenesis
8 pages, 4 figures, 3 tablesThe aim of this study was to
provide evidence on the modulation of lysosomal
enzymes in terms of both gene expression and
enzymatic activity during follicle maturation. For this
purpose three lysosomal enzymes, cathepsins B, D,
and L, were studied in relation to yolk formation and
degradation, during the main phases of ovarian follicle
growth in the pelagophil species, the sea bream
Sparus aurata. Specific attention was focused on the
gene expression quantification method, on the assay of
enzymatic activities, and on the relationship between
the proteolytic cleavage of yolk proteins (YPs), cathepsin
gene expression and cathepsin activities. For the
gene expression study, the cathepsins B-like and L-like
mRNAs were isolated and partially or fully characterized,
respectively; the sequences were used as design
specific primers for the quantification of cathepsin
gene expression by real-time PCR, in follicles at
different stages of maturation. The enzymatic assays
for cathepsins B, D, and L were optimized in terms of
specificity, sensitivity and reliability, using specific
substrates and inhibitors. In ovulated eggs, the lipovitellin
I (LV I) was degraded and the changes in
electrophoretic pattern were preceded by an increase in
the activity of a cysteine proteinase, cathepsin L, and
its mRNA. Cathepsin B did not appear to be involved
in YP changes during the final maturation stage.European Commission (DG Fisheries); Grant number:
Project #Q5RS-2002-00784-CRYOCYTE.Peer reviewe