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Abstract
Molecular determinants and mechanisms involved in ovarian
follicular growth, ovulation
and luteinization are not well understood. The objective
of this study was to identify genes
expressed in bovine granulosa cells (GC) of dominant
follicles (DF) and down-regulated after
hCG-induced ovulation, using the suppression subtractive
hybridization (SSH). GC were
collected from DF at day 5 of the estrous cycle and from
ovulatory follicles (OF) obtained 23 h
following injection of hCG. A subtracted cDNA library
(DF-OF) was generated and screened
using unsubtracted (DF, OF) and subtracted (DF-OF, OF-DF)
cDNAs as complex 32P-probes. A
total of 32 non-redundant cDNAs were identified: 23 cDNAs
matched with sequences of known
biological function and 9 cDNAs with complete or partial
sequences of undefined biological
function. Detection of genes known to be down-regulated
during the periovulatory period in the
bovine species, such as CPD, CYP11A1, CYP19A1, FSHR,
LRP8/ApoER2 and SERPINE2,
validated the physiological model and analytical
techniques used. For a subset of genes such as
ARFGAP3, CYP11A1, CYP19A1, FSHR, FST, GJA1, IDH3,
INHBA, LHCGR, LHCGR lacking
exon 10, PRC1, PRG1, RPA2, SCD and TRIB2, gene
expression profiles were compared by
virtual Northern blot or reverse transcriptase-polymerase
chain reaction from follicles obtained at
different developmental stages. Results confirmed a
down-regulation of the respective mRNAs in
GC of OF compared with that of DF. We conclude that we
have identified novel genes that are
down-regulated by hCG in bovine GC of DF during the
periovulatory period, which may
contribute to follicular growth, ovulation and/or
luteinization.
Key words:
Follicle •
Gene regulation •
Granulosa cells •
Ovulation
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