Subtractive hybridization reveals tissue‐specific expression of ahnak during embryonic development

The gene product ahnak has been identified from extra‐embryonic mesoderm cDNA enriched using a subtractive hybridization approach modified for using small amounts of starting material. Clones for cyclin D2 and H19 have also been isolated as being preferentially enriched in the extra‐embryonic mesoderm compared with the embryo proper of embryonic day (E) 7.5 neural plate stage mouse embryos. The differential expression of these genes was confirmed at gastrulation stage using in situ hybridization. More detailed analysis of the human genomic ahnak sequence suggests that its highly repetitive structure was formed by unequal cross‐over and gene conversion. During organogenesis, ahnak is expressed in a variety of tissues, including migratory mesenchyme. By E12.5, the major site of expression of ahnak is craniofacial mesenchyme. Immunohistochemical analysis has shown that ahnak protein is expressed mainly at the cell membrane of migratory mesenchymal cells, primarily in the nucleus of bone growth plate cells and mostly in the cytoplasm of differentiating nasal epithelia. The potential functions of ahnak are discussed in light of these results.