C. elegans heterochronic genes ensure that certain developmental events occur at the proper time. One such event is the terminal differentiation of lateral hypodermal seam cells which occurs during the final (L4) molt and is marked by cell cycle exit, fusion, and secretion of adult-specific cuticle containing lateral alae. The heterochronic gene
hbl-1 encodes a hunchback-like transcription factor.
hbl-1(
ve18) mutants have a precocious phenotype in which the seam cells fuse and secrete adult cuticle at the L3 molt. However, the seam nuclei divide again late in the L4 stage, resulting in seam syncytia with up to 12 extra nuclei. Complete depletion of
hbl-1 by RNAi produces an embryonic-lethal phenotype, whereas post-embryonic depletion results in a
hbl-1(
ve18)-like phenotype, indicating that
ve18 is a hypomorphic allele that produces sufficient
hbl-1 function for embryonic survival but not for timing post-embryonic development. Occasional skipping of the proliferative seam cell division at the L1 molt also occurs in post-embryonically depleted animals. Together, these results suggest
hbl-1 is required at several stages of development. Post-embryonic
hbl-1::gfp expression is temporally down-regulated in the hypodermis and nervous system in a 3'UTR-dependent manner. The
hbl-1 3'UTR contains multiple phylogenetically-conserved elements with complementarity to the
let-7 microRNA, and loss of
hbl-1 function suppresses the retarded heterochronic phenotype of
let-7, suggesting
hbl-1 is a
let-7 target.
hbl-1 is partially redundant with the heterochronic gene
lin-41, a
let-7 suppressor that is a known
let-7 target. To test whether
hbl-1 is regulated by
let-7,
hbl-1::gfp expression was compared between wild-type and
let-7 mutants. In
hyp7,
hbl-1::gfp expression is highest early in the L1 stage, drops dramatically by the L2, and is undetectable by the late L3 stage. This expression pattern is essentially unchanged in a
let-7 mutant background. Interestingly, in the seam,
hbl-1::gfp expression is not 3'UTR dependent and is detected only early in the L1, significantly prior to the L3 molt phenotype. One possibility is that
hbl-1 activity in
hyp7 regulates temporal identities in the seam; mosaic analysis will test this idea. In the nervous system,
hbl-1::gfp expression is also highest in the L1 stage but drops gradually, and is often not detectable in adults. Enhanced neuronal
hbl-1::gfp expression in late stages was seen in
let-7 mutants, indicating that
hbl-1 is regulated by
let-7 in this tissue. This modest deregulation of
hbl-1 in
let-7 mutants suggests that there may be redundant factors that time
hbl-1 down-regulation, perhaps including the recently identified
let-7 homologs.