A pathway of heterochronic genes act through
lin-29 to regulate the terminal differentiation of hypodermal cells (the 'L/A switch') at the L4 molt . Thus,
lin-29 gene product, a Zn-finger protein (see Rougvie and Ambros, these abstracts), likely regulates the stage-specific transcription of 'downstream' genes that include genes that control cell division, cell fusion, cuticle synthesis and molting. Using Northern blot analysis, we showed that the proper temporal regulation of certain C. elegans cuticle collagen genes depends upon
lin-29 activity. Since
lin-29 is required in continuous and post-dauer development, the downstream genes may also be shared by both pathways. In support of this, we have found that the steady-state transcript levels from certain collagen genes show the same stage-specificity during both continuous and post-dauer development. To further examine the regulation of larval and adult-specific collagen gene transcription by the heterochronic gene pathway, we inserted 5' upstream sequences of col-
l9 and
col-7 (adult-specific) and
col-17 ( larval specific) into lac-Z reporter vectors (developed by Andy Fire). These fusion constructs were injected into wild type and heterochronic mutant animals using the
rol-6(sulO06) PRF4 plasmid to identify rolling transformed lines containing extrachromosomal arrays of PRF4 and the fusion constructs. In wild type transformed lines, the
col19 and
col-7 fusions were expressed only in adults, beginning from L4 lethargus, and the
col-17 fusion was expressed from late embryogenesis to the L4 and not in adults. As little as 190 bp 5' of the col-l 9 translation start is sufficient for adult-specific expression. The expression of each construct in heterochronic mutants was consistent with col-l 9 and
col-7 being activated by
lin-29 at the L4 molt, and
col-17 being repressed by
lin-29 at the L4 molt. Experiments are underway to test whether the
lin29 product regulates any of these collagen genes directly. The
col-19llac-Z and
col-711ac-Z fusions show the same stagespecificity during both continuous and post-dauer development. Further, the precocious expression of
col-19llac-Z in lin- 14 animals is corrected by post-dauer development. Thus, the morphological suppression of precocious L/A switching in
lin-4,
lin-14 or
lin-28 animals during postdauer development likely reflects a restoration of the proper stage-specific transcription of col-l 9 and perhaps other downstream genes.