Biology subjects

Phillips, B. T.

Publications and source records attributed to Phillips, B. T..

3 recordsLinked to original sources

bioRxiv2022

unc-37/Groucho and lsy-22/AES repress Wnt target genes in C. elegans asymmetric cell divisions

Asymmetric cell division (ACD) is a fundamental mechanism of cell fate specification and adult tissue homeostasis. In C. elegans, the Wnt/{beta}-catenin asymmetry (W{beta}A) pathway regulates ACDs throughout embryonic and larval development. Under control of Wnt ligand-induced polarity, the transcription factor POP-1/TCF functions with the coactivator SYS-1/{beta}-catenin to activate gene expression in the signaled cell or, in absence of the coactivator, to repress Wnt target genes in the unsignaled daughter cell. To date, investigation of Groucho function in W{beta}A is lacking, and the function of LSY-22/AES has only been evaluated in C. elegans neurons. Further, conflicting evidence shows TCF utilizing Groucho-mediated repression may be either aided or repressed by AES addition. Here we demonstrate a genetic interaction between Groucho corepressors and POP-1/TCF in the distal tip cells (DTCs), seam cells (SCs) and embryonic endoderm development. In the DTCs, signaled cell fate increases after individual and double Groucho loss of function, representing the first demonstration of Groucho function in wildtype W{beta}A ACDs. Further, W{beta}A target gene misexpression occurs more frequently than DTC fate changes, suggesting derepression generates an intermediate cell fate. In the SCs, loss of UNC-37/Groucho or LSY-22/AES in a POP-1/TCF hypomorphic background enhances SC expansion and target gene misregulation. Moreover, while POP-1/TCF depletion in lsy-22/AES nulls yielded an expected increase in SCs we observed a surprising SC decrease in unc-37/Groucho nulls subjected to POP-1/TCF depletion. This phenotype correlates with UNC-37/Groucho regulation of pop-1/tcf expression since POP-1/TCF levels are increased in unc-37/Groucho null SCs. Lastly, Groucho functions in embryonic endoderm development since we observe ectopic endoderm transgene expression in unc-37/Groucho and lsy-22/AES knockdown in a HDA-1 background. Together, these data indicate Groucho-mediated modulation of cell fate via regulation of POP-1/TCF repression is widespread in W{beta}A ACDs and suggests a novel role of LSY-22/AES as a bona fide TCF repressor.

developmental biology↗

bioRxiv2021

Nuclear localization and transactivation of SYS-1/β-catenin is the result of serial gene duplications and subfunctionalizations

{beta}-catenin is a multifunctional protein capable of mediating cell adhesion via E-cadherin and transactivation of target genes of the canonical Wnt signaling pathway. The nematode, C. elegans contains four paralogs of {beta}-catenin which are highly specific in their functions. Though similar in overall structure, the four beta-catenins are functionally distinct, each regulating different aspects of development. Of the four, SYS-1 is a key player in Wnt dependent asymmetric cell division (ACD). In ACD, a polarized mother will give rise to a daughter with high nuclear SYS-1 and another with low nuclear SYS-1. Despite sequence dissimilarity, SYS-1 shares a close structural resemblance with human {beta}-catenin where it retains an unstructured amino-terminus (NTD) and 12 armadillo repeats. Using existing genome sequence data from several nematode species, we find that the four {beta}-catenin paralogs result from 3 sequential gene duplications and neofunctionalizations during nematode evolution. SYS-1, however, lacks an unstructured carboxyl-terminus (CTD) that is essential for human {beta}-catenin transactivation processes. This work supports the hypothesis that SYS-1 compensated for the lack of CTD by acquiring novel transactivation domains with cryptic nuclear localization signals in the NTD and the first four armadillo repeats, as shown by transactivation assays in worms and yeast. Furthermore, SYS-1 regulatory domains are not localized to the NTD as in canonical {beta}-catenin and instead spans the entire length of the protein. Truncating SYS-1 abolishes the classical SYS-1 nuclear asymmetry, resulting in daughter cells with symmetrical SYS-1 truncation localization. A screen for SYS-1 physical interactors followed by in vivo cell fate and SYS-1 localization analyses suggest that proper SYS-1 nuclear export is facilitated by XPO-1, while an interaction with IMB-3, an importin {beta}-like protein, suggests import mechanisms. Interestingly, XPO-1 is especially required for lowering SYS-1 in the Wnt-unsignaled nucleus, suggesting a distinct mechanism for regulating asymmetric nuclear SYS-1. In summary, we provide insights on the mechanism of {beta}-catenin evolution within nematodes and inform SYS-1 transactivation and nuclear transport.

developmental biology↗

bioRxiv2021

Centrosomal Enrichment and Proteasomal Degradation of SYS-1/-β-catenin Requires the Microtubule Motor Dynein

The C. elegans Wnt/{beta}-catenin Asymmetry (W{beta}A) pathway utilizes asymmetric regulation of SYS- 1/{beta}-catenin and POP-1/TCF coactivators. W{beta}A differentially regulates gene expression during cell fate decisions, specifically by asymmetric localization of determinants in mother cells to produce daughters biased towards their appropriate cell fate. Despite the induction of asymmetry, {beta}-catenin localizes symmetrically to mitotic centrosomes in both mammals and C. elegans. Due to the mitosis-specific localization of SYS-1 to centrosomes and enrichment of SYS-1 at kinetochore microtubules when SYS-1 centrosomal loading is disrupted, we investigated active trafficking in SYS-1 centrosomal localization. Here, we demonstrate that trafficking by microtubule motor dynein is required to maintain SYS-1 centrosomal enrichment, by dynein RNAi-mediated decreases in SYS-1 centrosomal enrichment and by temperature-sensitive allele of the dynein heavy chain. Conversely, we observe depletion of microtubules by nocodazole treatment or RNAi of dynein-proteasome adapter ECPS-1 exhibits increased centrosomal enrichment of SYS-1. Moreover, disruptions to SYS-1 or negative regulator microtubule trafficking are sufficient to significantly exacerbate SYS-1 dependent cell fate misspecifications. We propose a model whereby retrograde microtubule-mediated trafficking enables SYS-1 enrichment at centrosomes, enhancing its eventual proteasomal degradation. These studies support the link between centrosomal localization and enhancement of proteasomal degradation, particularly for proteins not generally considered centrosomal.

cell biology↗