Identification of evolutionarily conserved promoter elements and amino acids required for function of the C. elegans β-catenin homolog BAR-1

Abstract

beta-catenins are conserved transcription factors regulated posttranslationally by Wnt signaling. bar-1 encodes a Caenorhabditis elegans beta-catenin acting in multiple Wnt-mediated processes, including cell fate specification by vulval precursor cells (VPCs) and migration of the Q(L) neuroblast progeny. We took two approaches to extend our knowledge of bar-1 function. First, we undertook a bar-1 promoter analysis using transcriptional GFP reporter fusions and found that bar-1 expression is regulated in specific cells at the transcriptional level. We identified promoter elements necessary for bar-1 expression in several cell types, including a 321-bp element sufficient for expression in ventral cord neurons (VCNs) and a 1.1-kb element sufficient for expression in the developing vulva and adult seam cells. Expression of bar-1 from the 321-bp element rescued the Uncoordinated (Unc) phenotype of bar-1 mutants, but not the vulval phenotype, suggesting that a Wnt pathway may act in ventral cord neurons to mediate proper locomotion. By comparison of the 1.1-kb element to homologous sequences from Caenorhabditis briggsae, we identified evolutionarily conserved sequences necessary for expression in vulval or seam cells. Second, we analyzed 24 mutations in bar-1 and identified several residues required for BAR-1 activity in C. elegans. By phylogenetic comparison, we found that most of these residues are conserved and may identify amino acids necessary for beta-catenin function in all species.

Anchor cell signaling and vulval precursor cell positioning establish a reproducible spatial context during C. elegans vulval induction

“…lin-3(e1417) animals also showed a significantly reduced proportion of individuals with complete AC-P6.p alignment (25% compared to 76% in wild type) (Fisher's Exact Test, P<0.0001). In -1(ga80) is a putative null allele, while bar-1(mu63) is a weak allele (Natarajan et al, 2004). (C) Combination of mutants of the EGF/Ras/MAPK and Wnt signaling pathways (purple).…”

The β-catenin HMP-2 functions downstream of Src in parallel with the Wnt pathway in early embryogenesis of C. elegans

“…First, ectopic overexpression of HMP-2 rescues the mutant phenotype of bar-1 encoding another β-catenin that binds POP-1 and promotes its nuclear translocation and activation of transcriptional activity (Natarajan et al, 2004). Second, HMP-2 can substitute the endoderm inducing activity of yet another β-catenin WRM-1, which binds POP-1 to exclude POP-1 from the nucleus, in the absence of FRK-1 (Putzke and Rothman, 2010).…”

C. elegansGATA factors EGL-18 and ELT-6 function downstream of Wnt signaling to maintain the progenitor fate during larval asymmetric divisions of the seam cells

“…We expressed FLAGtagged polyA-binding protein (PAB-1) in the seam cells using a previouslydescribed bar-1 enhancer element (Natarajan et al, 2004). Synchronized L1 worms of control (N2) and experimental strains huIs1;deIs10 (for activation of Wnt signaling) and deIs26;deIs10 (for inhibition of Wnt signaling) were heat shocked at the L2/L3 molt as described previously (Gleason et al, 2002;Gleason and Eisenmann, 2010).…”

“…We have previously shown that seam cell fate specification is sensitive to alterations in Wnt signaling at this time (Gleason and Eisenmann, 2010). Transcripts expressed in the seam cells (and VPCs) were enriched by the mRNA-tagging method (Roy et al, 2002), in which we used a previously described bar-1 enhancer (Natarajan et al, 2004) to drive tagged polyA binding protein (PAB-1) expression in those cells. Seam/VPC-enriched transcript pools from experimental and control strains were analyzed using Affymetrix microarrays, and a 240 genes that were upregulated in the Wnt-activated samples compared with the Wnt-inhibited samples were identified (L.G., W. Chen, T. Brodigan, M. Krause and D.M.E., unpublished).…”