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BOR - Papers in Press, published online ahead of print September 1, 2004.
Biol Reprod 2004, 10.1095/biolreprod.104.029850
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BIOLOGY OF REPRODUCTION 72, 50–57 (2005)
DOI: 10.1095/biolreprod.104.029850
© 2005 by the Society for the Study of Reproduction, Inc.

Expression, Regulation, and Immunolocalization of Putative Homeodomain Transcription Factor 1 (PHTF1) in Rodent Epididymis: Evidence for a Novel Form Resulting from Proteolytic Cleavage1

J. Oyhenart3,4, J.L. Dacheux5, F. Dacheux5, B. Jégou6, and N. Raich2,4

INSERM U 567 CNRS-UMR 8104,4 Institut Cochin, Département d'Hématologie, Maternité de Port-Royal, Université René Descartes, 75014 Paris, France Station de Physiologie de la Reproduction et des Comportements,5 UMR INRA-CNRS 6073, 37380 Nouzilly, France GERM-INSERM U. 435 Université de Rennes I,6 Campus de Beaulieu, 35042 Rennes cedex, Bretagne, France

PHTF1 is an 84–86-kDa membrane protein found in the endoplasmic reticulum of male germ cells in rodents. There are no evident signs of PHTF1 in the spermatozoa released into the lumen of the seminiferous tubules but PHTF1 is present in the epididymal epithelium. Characterization of the epididymal Phtf1 messenger by Northern blot and reverse transcription-PCR identified a 3-kilobase transcript in the epididymis, similar to that previously reported in the testis. The transcript is present in the proximal part of the epididymis and it appears when the rats reach 4 wk of age. Through immunofluorescence analysis, PHTF1 was localized in the principal cells of the initial segment and the caput epididymis. Colocalization with different markers indicated PHTF1 is in the endoplasmic reticulum saccules applied to the trans face of the Golgi system. Western blot analyses revealed a shorter form of the protein—about 56-kDa versus the 84-kDa form found in the testis. Using the canine epididymal cell line CIM 20, transfected by N- and C-terminal myc-tagged PHTF1, we demonstrated that the 56-kDa epididymal form could result from proteolytical processing.

1 Supported by INSERM, INRA, and CNRS and by CONICET and FRM grants to J.O.

2 Correspondence: Raich Natacha, INSERM U 567-UMR 8104, 123 boulevard de Port-Royal, 75014 Paris, France. FAX: 33 014 325 1167; raich{at}infobiogen.fr

3 Current address: UCSF Comprehensive Cancer Center, 2340 Sutter Street, Box 0128, Room N-171, San Francisco, CA 94143-0128






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