Molecular cloning and analysis of a group of genes differentially expressed in cells which overexpress the Hoxa-1 homeobox gene

The homeobox gene Hoxa-1 is transcriptionally regulated by retinoic acid (R A) and encodes a transcription factor which has been shown to play importan t roles in cell differentiation and embryogenesis. In order to clone and ch aracterize target genes of Hoxa-1, we utilized differential hybridization s creening and cDNA subtractive hybridization methods to identify genes which are differentially expressed in F9-10, a murine F9 teratocarcinoma stem ce ll line which expresses high levels of exogenous Hoxa-1, compared to F9 wil d-type stem cells, which do not express endogenous Hoxa-1 mRNA in the absen ce of RA. Twenty-eight candidate genes were identified; these genes encode very diverse proteins, including signaling molecules such as BMP-4, the enz yme superoxide dismutase, the cell adhesion molecule cadherin-6, proteins i nvolved in gene transcription such as HMG-1 and SAP18, homeodomain-containi ng proteins Gbx-2 and Evx-8, and cell cycle regulatory proteins such as the retinoblastoma binding protein-a. Clone 104 encodes a novel protein; the e xpression of the clone 104 mRNA is also regulated in a fashion very similar to that of the exogenous Hoxa-1 gene in another F9 cell line, called F9-te t Hoxa1-8, in which the exogenous Hoxa-1 mRNA expression is tightly regulat ed by a Tet-off gene expression system. These data strongly suggest that cl one 104 is a direct downstream target of the transcription factor Hoxa-1. T he cDNA sequence of clone 104 is related to that of human ubiquitin carboxy l-terminal hydrolase T. Further characterization of these putative Hoxa-1 t arget genes will aid in delineating the functions of the Hoxa-1 protein in the differentiation processes which occur during embryogenesis. (C) 2000 Ac ademic Press.