Beta-catenin asymmetry is regulated by PLA1 and retrograde traffic in C. elegans stem cell divisions

Asymmetric division is an important property of stem cells. In Caenorhabditis elegans, the Wnt/beta-catenin asymmetry pathway determines the polarity of most asymmetric divisions. The Wnt signalling components such as beta-catenin localize asymmetrically to the cortex of mother cells to produce two distinct daughter cells. However, the molecular mechanism to polarize them remains to be elucidated. Here, we demonstrate that intracellular phospholipase A(1) (PLA(1)), a poorly characterized lipid-metabolizing enzyme, controls the subcellular localizations of beta-catenin in the terminal asymmetric divisions of epithelial stem cells (seam cells). In mutants of ipla-1, a single C. elegans PLA(1) gene, cortical beta-catenin is delocalized and the asymmetry of cell-fate specification is disrupted in the asymmetric divisions. ipla-1 mutant phenotypes are rescued by expression of ipla-1 in seam cells in a catalytic activity-dependent manner. Furthermore, our genetic screen utilizing ipla-1 mutants reveals that reduction of endosome-to-Golgi retrograde transport in seam cells restores normal subcellular localization of beta-catenin to ipla-1 mutants. We propose that membrane trafficking regulated by ipla-1 provides a mechanism to control the cortical asymmetry of beta-catenin.     

Crosstalk between a nuclear receptor and beta-catenin signaling decides cell fates in the C. elegans somatic gonad

beta-Catenin signaling determines the proximal-distal axis of the C. elegans gonad by promoting distal fate in asymmetrically dividing somatic gonad precursor cells (SGPs). Impaired function of the Wnt effector POP-1/TCF, its coactivator SYS-1/beta-catenin, and of upstream components including beta-catenin WRM-1 causes all SGP daughters to adopt the proximal fate. Consequently, no distal tip cells (DTCs) that would lead differentiation of gonad arms form in the affected hermaphrodites. Here, we show that deficiency of the nuclear receptor NHR-25 has the opposite effect: extra DTCs develop instead of proximal cells. NHR-25 knockdown restores DTC formation and fertility in pop-1 and sys-1 mutants, suggesting that a balance between NHR-25 and beta-catenin pathway activities is required to establish both proximal and distal fates. This balance relies on direct crossregulation between NHR-25 and the distinct beta-catenin proteins WRM-1 and SYS-1. The nuclear receptor-beta-catenin interaction may be an ancient mechanism of cell-fate decision.     

tcl-2 encodes a novel protein that acts synergistically with Wnt signaling pathways in C. elegans

Mutations in tcl-2 cause defects in the specification of the fates of the descendants of the TL and TR blast cells, whose polarity is regulated by lin-44/Wnt and lin-17/frizzled, during Caenorhabditis elegans development. In wild-type animals, POP-1/TCF/LEF, is asymmetrically distributed to the T cell daughters, resulting in a higher level of POP-1 in the nucleus of the anterior daughter. The POP-1 asymmetric distribution is controlled by lin-44 and lin-17. However, in tcl-2 mutants, POP-1 is equally distributed to T cell daughters as is observed in lin-17 mutants, indicating that, like lin-17, tcl-2 functions upstream of pop-1. In addition, tcl-2 mutations cause defects in the development of the gonad and the specification of fate of the posterior daughter of the P12 cell, both of which are controlled by the Wnt pathway. Double mutant analyses indicate that tcl-2 can act synergistically with the Wnt pathway to control gonad development as well as P12 descendant cell fate specification. tcl-2 encodes a novel protein. A functional tcl-2::gfp construct was weakly expressed in the nuclei of the T cell and its descendants. Our results suggest that tcl-2 functions with Wnt pathways to control T cell fate specification, gonad development, and P12 cell fate specification.     

DSH-2 regulates asymmetric cell division in the early C. elegans somatic gonad

Like other organs, the C. elegans gonad develops from a simple primordium that must undergo axial patterning to generate correct adult morphology. Proximal/distal (PD) polarity in the C. elegans gonad is established early during gonadogenesis by the somatic gonad precursor cells, Z1 and Z4. Z1 and Z4 each divide asymmetrically to generate one daughter with a proximal fate and one with a distal fate. PD polarity of the Z1/Z4 lineages requires the activity of a Wnt pathway that activates the TCF/LEF homolog pop-1. How the gonadal pathway controlling pop-1 is regulated by upstream factors has been unclear, as neither Wnt nor Dishevelled (Dsh) proteins have been shown to be required. Here we show that the C. elegansdsh homolog dsh-2 controls gonadal polarity. As in pop-1 mutants, dsh-2 hermaphrodites have Z1 and Z4 lineage defects indicative of defective PD polarity and are missing gonadal arms. Males have an elongated but disorganized gonad, also with lineage defects. DSH-2 protein is expressed in the Z1/Z4 gonadal precursor cells. Asymmetric distribution of nuclear GFP::POP-1 in Z1 and Z4 daughter cells is reversed in dsh-2 mutants, with higher levels in distal than proximal daughters. dsh-2 and the frizzled receptor homolog lin-17 have a strong genetic interaction, suggesting that they act in a common pathway. We suggest that DSH-2 functions as an upstream regulator of POP-1 in the somatic gonad to control asymmetric cell division, thereby establishing proximal-distal polarity of the developing organ.     

Control of cell polarity by noncanonical Wnt signaling in C. elegans

The three Caenorhabditis elegans beta-catenin each function in distinct processes: BAR-1 in canonical Wnt signaling that controls cell fates and cell migrations, HMP-2 in cell adhesion and WRM-1 in Wnt signaling pathways that function in conjunction with a mitogen-activated kinase (MAPK) pathway to control the orientations, or cell polarities, of cells that undergo asymmetric cell divisions. In addition, WRM-1 does not interact with the canonical beta-catenin binding site in POP-1/Tcf. Thus, Wnt signaling through WRM-1 is noncanonical and, except for one division that might not include any of the three C. elegans beta-catenin, controls cell polarity in C. elegans.     

A novel noncanonical Wnt pathway is involved in the regulation of the asymmetric B cell division in C. elegans

The polarities of several cells that divide asymmetrically during Caenorhabditis elegans development are controlled by Wnt signaling. LIN-44/Wnt and LIN-17/Fz control the polarities of cells in the tail of developing C. elegans larvae, including the male-specific blast cell, B, that divides asymmetrically to generate a larger anterior daughter and a smaller posterior daughter. We determined that WRM-1 and the major canonical Wnt pathway components: BAR-1, SGG-1/GSK-3 and PRY-1/Axin were not involved in the control of B cell polarity. However, POP-1/Tcf is involved and is asymmetrically distributed to the B daughter nuclei, as it is in many cell divisions during C. elegans development. Aspects of the B cell division are reminiscent of the divisions controlled by the planar cell polarity (PCP) pathway that has been described in both Drosophila and vertebrate systems. We identified C. elegans homologs of Wnt/PCP signaling components and have determined that many of them appear to be involved in the regulation of B cell polarity. Specifically, MIG-5/Dsh, RHO-1/RhoA and LET-502/ROCK appear to play major roles, while other PCP components appear to play minor roles. We conclude that a noncanonical Wnt pathway, which is different from other Wnt pathways in C. elegans, regulates B cell polarity.     

Wnt signaling and a Hox protein cooperatively regulate psa-3/Meis to determine daughter cell fate after asymmetric cell division in C. elegans

Asymmetric cell division is a mechanism for achieving cellular diversity. In C. elegans, many asymmetric cell divisions are controlled by the Wnt-MAPK pathway through POP-1/TCF. It is poorly understood, however, how POP-1 determines the specific fates of daughter cells. We found that nob-1/Hox, ceh-20/Pbx, and a Meis-related gene, psa-3, are required for asymmetric division of the T hypodermal cell. psa-3 expression was asymmetric between the T cell daughters, and it was regulated by POP-1 through a POP-1 binding site in the psa-3 gene. psa-3 expression was also regulated by NOB-1 and CEH-20 through a NOB-1 binding sequence in a psa-3 intron. PSA-3 can bind CEH-20 and function after the T cell division to promote the proper fate of the daughter cell. These results indicate that cooperation between Wnt signaling and a Hox protein functions to determine the specific fate of a daughter cell.     

Cortical beta-catenin and APC regulate asymmetric nuclear beta-catenin localization during asymmetric cell division in C. elegans

In C. elegans, Wnt signaling regulates a number of asymmetric cell divisions. During telophase, WRM-1/beta-catenin localizes asymmetrically to the anterior cortex and the posterior daughter's nucleus. However, cortical WRM-1's functions are not known. Here, we use a membrane-targeted form of WRM-1 to show that cortical WRM-1 inhibits Wnt signaling and the nuclear localization of WRM-1. These functions are mediated by APR-1/APC, which regulates WRM-1 nuclear export. We also show that APR-1 as well as PRY-1/Axin and Dishevelled homologs localize asymmetrically to the cortex. Our results suggest a model in which cortical WRM-1 recruits APR-1 to the anterior cortex before and during division, and the cortical APR-1 stimulates WRM-1 export from the anterior nucleus at telophase. Because beta-catenin and APC are localized to the cortex in many cell types in different species, our results suggest that these cortical proteins may regulate asymmetric divisions or Wnt signaling in other organisms as well.     

Wnt regulates spindle asymmetry to generate asymmetric nuclear β-catenin in C. elegans

Extrinsic signals received by a cell can induce remodeling of the cytoskeleton, but the downstream effects of cytoskeletal changes on gene expression have not been well studied. Here, we show that during telophase of an asymmetric division in C. elegans, extrinsic Wnt signaling modulates spindle structures through APR-1/APC, which in turn promotes asymmetrical nuclear localization of WRM-1/β-catenin and POP-1/TCF. APR-1 that localized asymmetrically along the cortex established asymmetric distribution of astral microtubules, with more microtubules found on the anterior side. Perturbation of the Wnt signaling pathway altered this microtubule asymmetry and led to changes in nuclear WRM-1 asymmetry, gene expression, and cell-fate determination. Direct manipulation of spindle asymmetry by laser irradiation altered the asymmetric distribution of nuclear WRM-1. Moreover, laser manipulation of the spindles rescued defects in nuclear POP-1 asymmetry in wnt mutants. Our results reveal a mechanism in which the nuclear localization of proteins is regulated through the modulation of microtubules.     

C. elegans POP-1/TCF functions in a canonical Wnt pathway that controls cell migration and in a noncanonical Wnt pathway that controls cell polarity

In Caenorhabditis elegans, Wnt signaling pathways are important in controlling cell polarity and cell migrations. In the embryo, a novel Wnt pathway functions through a (beta)-catenin homolog, WRM-1, to downregulate the levels of POP-1/Tcf in the posterior daughter of the EMS blastomere. The level of POP-1 is also lower in the posterior daughters of many anteroposterior asymmetric cell divisions during development. I have found that this is the case for of a pair of postembryonic blast cells in the tail. In wild-type animals, the level of POP-1 is lower in the posterior daughters of the two T cells, TL and TR. Furthermore, in lin-44/Wnt mutants, in which the polarities of the T cell divisions are frequently reversed, the level of POP-1 is frequently lower in the anterior daughters of the T cells. I have used a novel RNA-mediated interference technique to interfere specifically with pop-1 zygotic function and have determined that pop-1 is required for wild-type T cell polarity. Surprisingly, none of the three C. elegans (beta)-catenin homologs appeared to function with POP-1 to control T cell polarity. Wnt signaling by EGL-20/Wnt controls the migration of the descendants of the QL neuroblast by regulating the expression the Hox gene mab-5. Interfering with pop-1 zygotic function caused defects in the migration of the QL descendants that mimicked the defects in egl-20/Wnt mutants and blocked the expression of mab-5. This suggests that POP-1 functions in the canonical Wnt pathway to control QL descendant migration and in novel Wnt pathways to control EMS and T cell polarities.     

Wnt signaling controls the stem cell-like asymmetric division of the epithelial seam cells during C. elegans larval development

Metazoan stem cells repopulate tissues during adult life by dividing asymmetrically to generate another stem cell and a cell that terminally differentiates. Wnt signaling regulates the division pattern of stem cells in flies and vertebrates. While the short-lived nematode C. elegans has no adult somatic stem cells, the lateral epithelial seam cells divide in a stem cell-like manner in each larval stage, usually generating a posterior daughter that retains the seam cell fate and an anterior daughter that terminally differentiates. We show that while wild-type adult animals have 16 seam cells per side, animals with reduced function of the TCF homolog POP-1 have as many as 67 seam cells, and animals with reduced function of the β-catenins SYS-1 and WRM-1 have as few as three. Analysis of seam cell division patterns showed alterations in their stem cell-like divisions in the L2-L4 stages: reduced Wnt signaling caused both daughters to adopt non-seam fates, while activated Wnt signaling caused both daughters to adopt the seam fate. Therefore, our results indicate that Wnt signaling globally regulates the asymmetric, stem cell-like division of most or all somatic seam cells during C. elegans larval development, and that Wnt pathway regulation of stem cell-like behavior is conserved in nematodes.