Petratou, Dionysia et al. (2021) International Worm Meeting "The DEG/ΕNaC ion channel DEL-4 maintains neuronal ionstasis and promotes neuronal survival under stress"

Kaulich, Eva, Petratou, Dionysia, Gjikolaj, Martha, Schafer, William, Tavernarakis, Nektarios

[International Worm Meeting, 2021]

Physiological stress is sensed and processed by neurons, which initiate systemic stress responses. At the cellular level, ion homeostasis is of utmost importance for neuronal function, both for the maintenance of resting potential, and for the creation and propagation of action potentials. Indeed, imbalance in neuronal sodium homeostasis has been linked with many pathologies of the nervous system. However, the effects of stress on neuronal sodium homeostasis, excitability and survival are not understood. We find that DEL-4, an ENaC/DEG family member, which forms proton-inactivated homomeric sodium channels, is differentially regulated under stress to trigger appropriate cellular stress responses and motor adaptation. DEL-4 exhibits neuronal, non-synaptic localization and modulates the characteristics of Caenorhabditis elegans locomotory behavior. Heat stress and starvation alter DEL-4 expression, which in turn modulates the expression and activity of key stress response transcription factors, leading to increased autophagy and triggering of the ER stress response. Notably, similar to heat stress and starvation, DEL-4 deficiency causes hyperpolarization of dopaminergic neurons and impacts neurotransmission in dopaminergic and motor neurons. Utilizing two humanized models of Parkinson's and Alzheimer's disease in C. elegans, we show that DEL-4 promotes neuronal survival in the context of these proteinopathies. Our findings provide insight on the molecular mechanisms via which sodium channels uphold neuronal function and promote adaptation upon stress.

Kuniaki Nakamura et al. (2002) East Coast Worm Meeting "Isolation and characterization of the cks-1 mutant in C. elegans"

Kuniaki Nakamura, Craig C Mello, Soyoung Kim

[East Coast Worm Meeting, 2002]

In wild-type embryos the 4-cell stage blastomere, EMS, undergoes an inductive interaction that specifies the endoderm and orients the division axis of the EMS cell onto the anterior/posterior (a/p) axis of the embryo. In our temperature-sensitive mutant screen, we isolated one recessive mutant, ne549, with a penetrant EMS division orientation defect and ectopic endoderm produced by the C blastomere. A mutant with a very similar phenotype, cdk-1(ne236) was identified in a previous screen by Martha Soto (See ECWM abstract). We mapped ne549 to the right of unc-24 on LGIV, into a region that contains a C. elegans homolog of the highly conserved CDK-1 interacting protein, CKS-1, which encodes a conserved 13 kD protein required for cyclin-dependent kinase activity. We obtained rescue of ne549 with a single gene PCR product that contains the cks-1(+) allele, and we found a single point mutation in a highly conserved residue within the open reading of cks-1. Thus we conclude that ne549 is an allele of cks-1. RNAi of cks-1 causes a drastic cell division defect during early embryogenesis, therefore ne549 is clearly a special allele that alters but does not abolish the function of CKS-1. Given the similarity between cks-1(ne549) and cdk-1(ne236) phenotypes, we hypothesize that the two proteins function together in an activity that is required for endoderm specification and the control of division orientation. Although each single mutant exhibits wild-type cell-cycles we found that double mutants between cks-1(ne549) and cdk-1(ne236), resemble the null (RNAi) phenotypes for the individual mutants. This finding suggests that both mutants impair a function(s) of the kinase complex that are required for the cell-cycle. How do cell-cycle molecule(s) function to control both the orientation of the cell division and cell fate? This is a very interesting question but has yet to be answered in any organism. As one potential avenue to address the targets or interactors of CKS-1, we have started a suppressor screen for cks-1 mutants. So far we have identified 50 dominant suppressors, which arise at knock-out frequency and define at least three loci. We will report on our progress toward mapping, and characterizing these suppressors, as well as our further analysis of the cks-1(ne549) mutant.

Kuniaki Nakamura et al. (2003) International Worm Meeting "Characterization of the cks-1 mutant and its suppressor in C. elegans"

Kuniaki Nakamura, Soyoung Kim, Craig C Mello

[International Worm Meeting, 2003]

CKS-1 is a highly conserved cell cycle regulatory protein that physically associates with CDK-1 and is required for the full activation of CDK-1. In yeast, disruption of the cks-1 gene results in G1 arrest similar to that caused by mutants in cdc-28 (cdk-1). CKS-1 is required for the degradation of the cyclin dependent kinase inhibitor p27 in late G1 phase, allowing CDK activity to drive cells into S phase. Thus cks-1 plays a critical role in cell cycle progression. In our temperature-sensitive mutant screen, we isolated one recessive mutant, ne549, with a penetrant EMS division orientation defect and ectopic endoderm produced by the C blastomere. A mutant with a very similar phenotype, cdk-1(ne236) was identified in a previous screen in the lab by Martha Soto. We identified ne549 as an allele of cks-1 based on rescue with a single gene PCR product and we identified a single point mutation in a highly conserved residue within the open reading of cks-1. Interestingly, this lesion is a very subtle change that based on the crystal structure removes a single hydroxyl group from the protein and hence breaks a single intramolecular hydrogen bond. RNAi of cdk-1 or cks-1 causes a one-cell arrest phenotype, therefore ne549 is clearly a special allele that alters, but does not abolish, the function of CKS-1. Given the similarity between cks-1(ne549) and cdk-1(ne236) phenotypes, we hypothesize that the two proteins function together in an activity that is required for endoderm specification and the control of division orientation. Although each single mutant exhibits wild-type cell-cycles we found that double mutants between cks-1(ne549) and cdk-1(ne236), resemble the null (RNAi) phenotypes for the individual mutants. This finding suggests that both mutants impair an ability of the kinase complex to regulate both cell cycle and cell polarity. How do cell cycle molecule(s) function to control both the orientation of the cell division and cell fate? This is a very interesting question but has yet to be answered in any organism. As one potential avenue to address the targets or interactors of CKS-1, we have started a suppressor screen for cks-1 mutants. So far we have identified several dominant suppressors, including at least one that also appears to suppress cdk-1(ne236)). We will report on our progress toward elucidating the function of CKS-1 and its partner CDK-1 in controlling division orientation.