Expression pattern of chick achaete-scute homologue (CASH) associated with mid-embryogenesis (stages 23 to 28)

Using the conserved basic helix-loop-helix (bHLH) domain of AS-C and MASH, we amplified by polymerase chain reaction, the HLH region of a chick AS-C/MASH homologue from cDNA prepared from poly (A+) RNA isolated from a region rich in cardiac neural crest cells. A full-length cDNA clone was obtained from a stage 23 chick head cDNA library using this conserved sequence as a probe. Comparison of this clone with MASH1 revealed strong homologies in the bHLH domain and carboxyl terminal region of the conceptually translated protein. The clone was named chick achaete-scute homologue (CASH). This transcript has a single open reading frame that codes for a protein of 218 residues. The deduced amino acid sequence of CASH is 100% identical to MASH1 in the bHLH region and 83.9% identical over the entire protein. The bHLH domain of CASH is highly conserved within the AS-C family. Northern analysis indicates that CASH is expressed transiently in a defined spatial pattern. The highest expression is in the head between stages 23 and 26. Whole mount in situ hybridization suggests that CASH is expressed at stage 23 in the dorsal ventricular zone and central sensory pathways and later in the ventral ventricular zone and ventral horn. By stage 28, it is expressed ubiquitously in the peripheral nervous system including all of the neurons derived from the the neural crest and epipharyngeal placodes, as well as neural crest-derived chromaffin cells in the adrenal medulla. Amplification by polymerase chain reaction indicates that CASH is expressed in the cardiac neural fold and continues to be expressed in cultured cardiac neural crest cells.