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Molecular and Cellular Biology, August 1999, p. 5823-5832, Vol. 19, No. 8
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

A Novel 14-Base-Pair Regulatory Element Is Essential for In Vivo Expression of Murine beta 4-Galactosyltransferase-I in Late Pachytene Spermatocytes and Round Spermatids

Martin Charron,1 Nancy L. Shaper,1 Bhanu Rajput,1,dagger and Joel H. Shaper1,2,*

The Cell Structure and Function Laboratory, The Oncology Center,1 and Department of Pharmacology and Molecular Sciences,2 The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287-8937

Received 18 February 1999/Returned for modification 5 April 1999/Accepted 17 May 1999

During murine spermatogenesis, beginning in late pachytene spermatocytes, the beta 4-galactosyltransferase-I (beta 4GalT-I) gene is transcribed from a male germ cell-specific start site. We had shown previously that a 796-bp genomic fragment that flanks the germ cell start site and contains two putative CRE (cyclic AMP-responsive element)-like motifs directs correct male germ cell expression of the beta -galactosidase reporter gene in late pachytene spermatocytes and round spermatids of transgenic mice (N. L. Shaper, A. Harduin-Lepers, and J. H. Shaper, J. Biol. Chem. 269:25165-25171, 1994). We now report that in vivo expression of beta 4GalT-I in developing male germ cells requires an essential and previously undescribed 14-bp regulatory element (5'-GCCGGTTTCCTAGA-3') that is distinct from the two CRE-like sequences. This cis element is located 16 bp upstream of the germ cell-specific start site and binds a male germ cell protein that we have termed TASS-1 (transcriptional activator in late pachytene spermatocytes and round spermatids 1). The presence of the Ets signature binding motif 5'-GGAA-3' on the bottom strand of the TASS-1 sequence (underlined sequence) suggests that TASS-1 is a novel member of the Ets family of transcription factors. Additional transgenic analyses established that an 87-bp genomic fragment containing the TASS-1 regulatory element was sufficient for correct germ cell-specific expression of the beta -galactosidase reporter gene. Furthermore, when the TASS-1 motif was mutated by transversion, within the context of the original 796-bp fragment, transgene expression was reduced 12- to 35-fold in vivo.

* Corresponding author. Mailing address: Johns Hopkins University School of Medicine Oncology Center, Rm. 1-127, 600 North Wolfe St., Baltimore, MD 21287-8937. Phone: (410) 955-8879. Fax: (410) 502-5499. E-mail: jshaper{at}jhmi.edu.

dagger Present address: Osiris Therapeutics, Inc., Baltimore, MD 21231-3043.

Molecular and Cellular Biology, August 1999, p. 5823-5832, Vol. 19, No. 8
0270-7306/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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