A somatic proteoglycan controls Notch-directed germ cell fate

Communication between the soma and germline optimizes germ cell fate programs. Notch receptors are key determinants of germ cell fate but how somatic signals direct Notch-dependent germ cell behavior is undefined. Here we demonstrate that SDN-1 (syndecan-1), a somatic transmembrane proteoglycan, controls expression of the GLP-1 (germline proliferation-1) Notch receptor in the Caenorhabditis elegans germline. We find that SDN-1 control of a somatic TRP calcium channel governs calcium-dependent binding of an AP-2 transcription factor (APTF-2) to the glp-1 promoter. Hence, SDN-1 signaling promotes GLP-1 expression and mitotic germ cell fate. Together, these data reveal SDN-1 as a putative communication nexus between the germline and its somatic environment to control germ cell fate decisions.

A Limited and Diverse Set of Suppressor Mutations Restore Function to INX-8 Mutant Hemichannels in the Caenorhabditis elegans Somatic Gonad

In Caenorhabditis elegans, gap junctions couple cells of the somatic gonad with the germline to support germ cell proliferation and gametogenesis. A strong loss-of-function mutation (T239I) affects the second extracellular loop (EL2) of the somatic INX-8 hemichannel subunit. These mutant hemichannels form non-functional gap junctions with germline-expressed innexins. We conducted a genetic screen for suppressor mutations that restore germ cell proliferation in the T239I mutant background and isolated seven intragenic mutations, located in diverse domains of INX-8 but not the EL domains. These second-site mutations compensate for the original channel defect to varying degrees, from nearly complete wild-type rescue, to partial rescue of germline proliferation. One suppressor mutation (E350K) supports the innexin cryo-EM structural model that the channel pore opening is surrounded by a cytoplasmic dome. Two suppressor mutations (S9L and I36N) may form leaky channels that support germline proliferation but cause the demise of somatic sheath cells. Phenotypic analyses of three of the suppressors reveal an equivalency in the rescue of germline proliferation and comparable delays in gametogenesis but a graded rescue of fertility. The mutations described here may be useful for elucidating the biochemical pathways that produce the active biomolecules transiting through soma–germline gap junctions.

Mutations that Affect Channel Opening of Innexin Hemichannels in the C. elegans Gonad

In C. elegans, gap junctions couple cells of the somatic gonad with the germline to support germ cell proliferation and gametogenesis. We previously characterized a strong loss-of-function mutation (T239I) affecting the second extracellular loop (EL2) of the somatic INX-8 hemichannel subunit. These mutant hemichannels form non-functional gap junctions with germline-expressed innexins. Here we describe the characterization of mutations that restore germ cell proliferation in the T239I EL2 mutant background. We recovered seven intragenic mutations located in diverse domains of INX-8 but not the EL domains. These second-site mutations compensate for the original channel defect to varying degrees, from nearly complete wild-type rescue, to partial rescue of germline proliferation. One suppressor mutation (E350K) supports the innexin cryo-EM structural model that the channel pore opening is surrounded by a cytoplasmic dome. Two suppressor mutations (S9L and I36N) may form leaky hemichannels that support germline proliferation but cause the demise of somatic sheath cells. Phenotypic analyses of three other suppressors reveal an equivalency in the rescue of germline proliferation and comparable delays in gametogenesis but a graded rescue of fertility. These latter mutations may be useful to probe interactions with the biochemical pathways that produce the molecules transiting through soma-germline gap junctions.

Oocyte Aging is Controlled by Mitogen Activated Protein Kinase Signaling

Oogenesis is one of the first processes to fail during aging. In women, most oocytes cannot successfully complete meiotic divisions during the fourth decade of life. Studies of the nematode Caenorhabditis elegans have uncovered conserved genetic pathways that control lifespan, but our knowledge regarding reproductive aging in worms and humans is limited. Specifically, little is known about germline internal signals that dictate the oogonial biological clock. Here, we report a thorough characterization of the changes in the worm germline during aging. We found that shortly after ovulation halts, germline proliferation declines, while apoptosis continues, leading to a gradual reduction in germ-cell numbers. In late aging stages, we observed that meiotic progression is disturbed and crossover designation and DNA double-strand break repair decrease. In addition, we detected a decline in the quality of mature oocytes during aging, as reflected by decreasing size and elongation of interhomolog distance, a phenotype also observed in human oocytes. Many of these altered processes were previously attributed to MAPK signaling variations in young worms. In support of this, we observed changes in activation dynamics of MPK-1 during aging. We therefore tested the hypothesis that MAPK controls oocyte quality in aged worms using both genetic and pharmacological tools. We found that in mutants with high levels of activated MPK-1, oocyte quality deteriorates more rapidly than in wild-type worms, whereas reduction of MPK-1 levels enhances quality. Thus, our data indicate that MAPK signaling controls germline aging and could be used to attenuate the rate of oogenesis quality decline.

Non-autonomous regulation of germline stem cell proliferation by somatic MPK-1/MAPK activity in C. elegans

Extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) is a major positive regulator of cell proliferation that is often upregulated in cancer. Yet few studies have addressed ERK/MAPK regulation of proliferation within a complete organism. The C. elegans ERK/MAPK ortholog MPK-1 is best known for its control of somatic organogenesis and germline differentiation, but it also stimulates germline stem cell proliferation. Here we identify tissue-specific MPK-1 isoforms and characterize their distinct roles in germline function. The germline-specific MPK-1B isoform promotes germline differentiation, but has no apparent role in germline stem cell proliferation. By contrast, the soma-specific MPK-1A isoform promotes germline proliferation non-autonomously. Indeed, MPK-1A functions in the intestine or somatic gonad to promote germline proliferation, independently of its other known roles. We propose that a non-autonomous role of ERK/MAPK in stem cell proliferation may be conserved across species and other tissue types, with major clinical implications for cancer and other diseases.

Notch-Directed Germ Cell Proliferation Is Mediated by Proteoglycan-Dependent Transcription

Notch receptors are essential membrane-bound regulators of cell proliferation and differentiation in metazoa. In the nematode Caenorhabditis elegans, correct expression of GLP-1 (germline proliferation-1), a germline-expressed Notch receptor, is important for germ cell maintenance. However, mechanisms that regulate GLP-1 expression are undefined. Here, we demonstrate that an AP-2 transcription factor (APTF-2) regulates GLP-1 expression through calcium-dependent binding to a conserved motif in the glp-1 promoter. Our data reveals that SDN-1 (syndecan-1), a transmembrane proteoglycan, regulates a TRP calcium channel in the soma to modulate the interaction between APTF-2 and glp-1 promoter - thus providing a potential communication nexus between the germline and its somatic environment to control germ cell fate decisions.

VRK-1 extends life span by activation of AMPK via phosphorylation

Vaccinia virus–related kinase (VRK) is an evolutionarily conserved nuclear protein kinase. VRK-1, the single Caenorhabditis elegans VRK ortholog, functions in cell division and germline proliferation. However, the role of VRK-1 in postmitotic cells and adult life span remains unknown. Here, we show that VRK-1 increases organismal longevity by activating the cellular energy sensor, AMP-activated protein kinase (AMPK), via direct phosphorylation. We found that overexpression of vrk-1 in the soma of adult C. elegans increased life span and, conversely, inhibition of vrk-1 decreased life span. In addition, vrk-1 was required for longevity conferred by mutations that inhibit C. elegans mitochondrial respiration, which requires AMPK. VRK-1 directly phosphorylated and up-regulated AMPK in both C. elegans and cultured human cells. Thus, our data show that the somatic nuclear kinase, VRK-1, promotes longevity through AMPK activation, and this function appears to be conserved between C. elegans and humans.