GENOMICS 8,586~590 (1990) SHORT COMMUNICATION The Pit4 Transcription Factor Gene Is a Candidate for the Murine Snell Dwarf Mutation SALLY A. CAMPER,*,’ THOMAS L. SAUNDERS,* RONALD W. KATZ,* AND ROGER H. REEvEst *Department of Human Genetics, University of Michigan Medical School, Ann Arbor, Michigan 48 109-0618; and t Developmental Genetics Laboratory, Department of Physiology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 Received April 20, 1990; revised June 14, 1990 Two nonallelic mouse mutations with severe dwarf phe- notypes are characterized by a lack of growth hormone, prolactin, and thyroid stimulating hormone. The cells that normally synthesize these pituitary hormones express a common transcription factor called GHF- 1 or Pit- 1. Using an intersubspecific backcross, we have demonstrated tight linkage of the Pit-2 and Snell dwarf (dto) genes on mouse chromosome 16. No recombination was observed between Pit-l and dw in 110 individuals examined. Southern blot analysis of genomic DNA reveals that the Pit-l gene is rearranged in C3H/HeJ-d&/dw mice but not in coisogenic +/+ animals, providing molecular evidence that a lesion in the Pit-l gene results in the Snell dwarf phenotype. Dem- onstration of low levels of Pit-l expression in Ames dwarf (df) mice implies that both Pit-l and df expression may be required for pituitary differentiation. o 1990 Academic Press. Inc. Two nonallelic, autosomal recessive mutations in mice are characterized by pituitary defects leading to severe dwarfism. Mice homozygous for the Snell (dw) and Ames (df) dwarf mutations are indistinguishable from heterozygotes at birth, but adult homozygous animals are about one-third normal size. Both mu- tants are characterized by diminished anterior pitu- itary function. Growth hormone (GH), prolactin (PRL), and thyroid stimulating hormone (TSH) are not detectable, nor are their corresponding cell types, somatotrophs, lactotrophs, and thyrotrophs (Bartke, 1964,1979; Roux et al., 1982; Cheng et al., 1983). The lack of detectable PRL and GH at any time during development indicates that both mutations result in a failure to initiate synthesis of these hormones and suggests that these mutants provide models of 1 To whom correspondence should be addressed. blocked differentiation (Slabaugh et al., 1982). A transcription factor that is normally found in the three cell types that are absent from both dwarf mu- tants has been isolated. This transcription factor, called Pit-l or GHF-1, was cloned on the basis of spe- cific binding to the promoters of the GH and PRL genes and the PRL enhancer (Bodner et al., 1988; In- graham et aZ., 1988). A variety of functional assays have shown that Pit-l plays a role in transcription of the GH and PRL genes, although the importance of Pit-l for prolactin transcription is still controversial (Ingraham et aZ., 1988; Mangalam et aZ., 1989; Theill et al., 1989; Karin et aZ., 1990; Dolle et al., 1990). It is not clear whether Pit-l is involved in transcriptional activation of the 6 subunit of TSH, but it is found in some thyrotrophs (Ingraham et aZ.,1988; Alexander et al., 1989). The factor is an excellent candidate for one of the dwarf lesions because it is specific to the three cell types affected in the df and dw dwarfs, and it is known to be a transcription factor for two of the three missing hormones. A mouse Pit-l cDNA clone was prepared by reverse transcription of pituitary mRNA followed bypolymer- ase chain reaction (PCR) amplification (Krug and Berger, 1987; Saiki et al., 1988). Poly(A)+mRNA was prepared from pituitaries of C57BL/6J X SJL/J F2 mice, and oligo(dT)-primed synthesis of cDNA using reverse transcriptase was performed. Oligonucleo- tides for PCR were synthesized on the basis of the rat Pit-l nucleotide sequence (Ingraham et al., 1988) us- ing regions of Pit-l that are conserved between rat and bovine (Bodner et al., 1988) (nucleotides 139-167 and 778-807) with the incorporation of Hind111 and BamHI linkers to facilitate subsequent cloning. The 689-bp amplification product, corresponding to amino acid residues 47-269, was cloned into the Hind111 and BamHI sites of pBluescript KS- (Strata- osss-7543/90 $3.00 Copyright 0 1990 by Academic Press, Inc. All rights of reproduction in any form reserved. 586