GENE REGULATION AND FUNCTION - B
Wednesday, August 4, 2004
10:30 AM–12:30 PM
(687) CHARACTERIZATION OF cDNA LIBRARIES DEVELOPED FROM BOVINE ZONA PELLUCIDA AND CUMULUS CELLS.
Hamel, Mélanie1, Dufort, Isabelle1, Robert, Claude 1, Sirard, Marc-Andre1, 1 Université Laval, Québec, QC, Canada
ABSTRACT- Cumulus cells play an important role during oocyte development by supplying essential nutrients and/or messenger molecules to the oocyte. There are multiple forms of cell-cell communication between the oocyte and cumulus cells, including direct cell contact-mediated gap junctions in the zona pellucida. We thought it is maybe possible that mRNAs were supplied from cumulus cells to oocyte by transiting through the gap junction of the zona pellucida. To verify this hypothesis, two partial plasmid cDNA libraries were generated by using a SMART-PCR based amplification of reverse-transcribed mRNAs from isolated bovine zona pellucida or bovine cumulus cells. Each library was characterized as to the number of clones and average insert size. Over 300 sequencing attempts were performed for partial zona pellucida library and over 100 for cumulus cells partial library. Analysis of the sequences by clustering algorithms revealed 197 and 56 unique sequences for zona pellucida and cumulus cells partial library respectively. For the partial zona pellucida library, 40% of total genes found were known whereas for the partial cumulus cells library, 61% of total genes found were known, showing that sequences in cumulus cells were more characterized than for zona pellucida. Furthermore, for known sequences, 50% were found from human genes and 38% from bovine genes for the zona pellucida and 15% of known sequences were from human genes and 85% from bovine genes for cumulus cells, another indication that bovine zona pellucida transcript are less known than cumulus cells genes. These results show that it might maybe possible that cumulus cells supply mRNAs to the oocyte by transiting through the gap junction of the zona pellucida or that translation could occurs closer to the oocyte membrane, but further experimentations are necessary to verify these possibility. The high percentage of unknown gene in zona pellucida can be explained by the fact that these genes are not highly expressed in cumulus cells. These genes could possibly constituted a sub-family since their localization indicate a potential segregation.
KEY WORDS: Oocyte, Zona pellucida, cDNA library, Cumulus cells