PARENT SESSION
SLIDE SESSION 11: GENE REGULATION & FUNCTION
Chairs: Michael Pallidino, Jeanne Wison-Rawles, Leslie Skalla (Trainee)
Univ Ottawa-Arts Hall 257
1:30 PM-3:30 PM

ECTOPIC EXPRESSION OF A SERTOLI-SPECIFIC HOMEOBOX GENE INCREASES DNA-STRAND BREAKS IN ADJACENT GERM CELLS.

Wayne, Chad¹, Sutton, Keith¹, Wilkinson, Miles¹, ¹

ABSTRACT- Homeobox genes encode a family of transcription factors that have been shown to regulate a number of developmental events including the specification of axial identity and organogenesis. Although much is known about the developmental role of homeobox genes during embryogenesis, we know little of their role in post-natal and adult events, including spermatogenesis, the process of germ cell maturation that occurs within the seminiferous tubules of the testis. The Pem homeobox gene is a candidate that may regulate spermatogenesis, as Pem is expressed in a stage-specific manner in Sertoli cells of the testes. To examine Pem's functional role in the testis, we developed transgenic mice that ectopically express Pem in Sertoli cells during all stages of the seminiferous epithelial cycle. Analysis of testis sections from transgenic and wild-type mice by TdT-mediated dUTP-biotin nick-end labeling (TUNEL) demonstrated that ectopic Pem expression resulted in an increase in internucleosomal DNA fragmentation in germ cells, but not in an increase in the number of apoptotic germ cells. We found that Pem increased DNA-strand breaks in both a cell type- and stage-specific manner. Preleptotene spermatocytes, round spermatids, and elongating spermatids (step 8-10) exhibited a large increase in TUNEL positivity in response to Pem-expressing Sertoli cells, whereas spermatogonia, pachytene spermatocytes, elongating spermatids (step 11+) and condensed spermatids had little or no change in TUNEL positivity. Analysis by the Comet assay demonstrated an increase in tail moment and in the percentage of DNA in the tails of testicular cells from mice ectopically expressing Pem. Studies are ongoing to determine the molecular mechanism for this effect. Our in vivo observations suggest that Pem is a transcription factor that regulates downstream target genes that have a role in mediating germ cell maturation events in the testis including, perhaps, the regulation of the chromatin packaging machinery.

KEY WORDS: Homeobox gene, Sertoli cell, DNA-strand breaks