Kersey, Rossio K. et al. (2011) International Worm Meeting "Transcriptional control of dorsal-ventral polarity."

Kersey, Rossio K., Fukushige, Tetsu, Brodigan, Thomas, Krause, Michael

[International Worm Meeting, 2011]

Spatial polarity cues in animals are used repeatedly during development for many processes, including cell fate determination, cell migration, and axon guidance. The body wall muscle cells of C. elegans are a great source of polarity cues because they extend the length of the animal in four quadrants, occupying left and right positions on both the dorsal and ventral side of the animal. One cue known to originate from body wall muscle is UNC-129/TGF-beta that is produced at higher levels in dorsal versus ventral muscle resulting in a dorsal-ventral gradient. This pattern of unc-129 expression requires the activity of the transcriptional repressor UNC-130 that is preferentially produced in ventral body wall muscle. To understand how these dorsal ventral differences in expression are established and maintained, we have analyzed the regulation of unc-130 expression and the distribution of UNC-130 protein. We found cis-acting elements in the promoter that drive ventral body wall muscle expression when isolated and that are sufficient to impose ventral polarity on the unc-54 promoter that normally is expressed uniformly in all body wall muscle. We have also defined the temporal distribution of UNC-130 in body wall muscle cells during embryogenesis. Our results clarify the mechanisms by which unc-130 regulates unc-129 expression and contributes to our understanding of the logic behind the generation of polarity cues.

Kersey, Rossio K. et al. (2009) International Worm Meeting "Transcriptional control of dorsal-ventral polarity in C. elegans."

Brodigan, Thomas, Kersey, Rossio K., Krause, Mike, Fukushige, Tetsu

[International Worm Meeting, 2009]

Spatial polarity cues in animals are used repeatedly during development for many processes, including cell fate determination, cell migration, and axon guidance. Once formed, the body wall muscle cells of C. elegans are a great source of polarity cues because they extend the length of the animal in four quadrants, occupying left and right positions on both the dorsal and ventral side of the animal. One cue known to originate from body wall muscle is UNC-129/TGF-beta that is produced at higher levels in dorsal versus ventral muscle resulting in a dorsal-ventral gradient. This pattern of unc-129 expression is, in turn, generated by the activity of the transcriptional repressor UNC-130 that is preferentially produced in ventral body wall muscle. To understand how these dorsal ventral differences in expression are established, we have analyzed the 10.5 Kb promoter region of unc-130 with a series of deletions. We found two regions in the promoter that drive ventral body wall muscle expression when isolated. These two promoter regions are also sufficient to impose ventral polarity on the unc-54 promoter that normally is expressed uniformly in all body wall muscle. Results from the present and future studies will contribute to elucidating the mechanisms of transcriptional repression by unc-130. By identifying the trans-acting factors regulating unc-130 expression we hope to understand the logic behind the generation of polarity cues.

Rossio K. Kersey et al. (2007) International Worm Meeting "Characterization of candidate transcription factors regulating muscle cell fate determination in Caenorhabditis elegans."

Tetsu Fukushige, Mike Krause, Rossio K. Kersey

[International Worm Meeting, 2007]

We have recently shown that ectopic HLH-1 and/or PAL-1 are able to convert all early somatic blastomeres to a muscle-like cell fate. We have used expression microarray profiling of embryonic muscle induced by these factors to identify additional factors acting in concert with HLH-1 to regulate myogenesis. Total RNA obtained from synchronized embryos collected at 2-hour intervals throughout a 6-hour period of development was hybridized to Affymetrix arrays. Thirty candidate transcription factors were found to be significantly induced in both HLH-1 and PAL-1 datasets. The present study has focused on two of these: the MADS-box transcription factor unc-120/SRF and the forkhead transcription factor unc-130. Complementary studies performed in our lab demonstrated that unc-120, in conjunction with hlh-1 and hnd-1, plays an essential role in body wall muscle differentiation. unc-120 is also expressed in other muscles throughout development and we are interested in understanding additional functions of UNC-120. Interestingly, we found that unc-120, hnd-1 double mutants showed a severe gonad/sterility phenotype. Our hypothesis is that UNC-120 may be required for gonadal sheath cell development, and the failure of those muscle-like cells to develop properly may explain why the germ cells within the defective gonad arms fail to development properly. The possible role of unc-120 in gonad development will be discussed. Previous studies done by Nash et al. (2000) showed that unc-130 is expressed only in ventral body wall muscle, where it acts as a repressor of unc-129 expression. This activity results in a reciprocal patter of expression, with unc-129 and unc-130 present only in the dorsal and ventral body wall muscle, respectively. This asymmetric pattern of expression, coupled with it presence in our microarray data, led to our interest in unc-130. The two main factors required for body wall muscle gene expression, hlh-1 and unc-120, are expressed equally in all body wall muscles, suggesting that other factors must be responsible for the asymmetric expression of unc-130. We hypothesize that activation of unc-130 in dorsal body wall muscle is repressed by an unknown, trans-acting regulator functioning through cis-acting gene regulatory elements. Preliminary results obtained from our tests of this model will be discussed.

Chun-Yi Huang et al. (2007) International Worm Meeting "eif-3.K is a pro-apoptotic gene in C. elegans."

Jia-Yun Chen, Yi-Chun Wu, Chun-Yi Huang

[International Worm Meeting, 2007]

Programmed cell death (or apoptosis) is an important feature of C. elegans development. Previous studies have identified pro-apoptotic genes egl-1, ced-3 and ced-4 and anti-apoptotic genes ced-9 and icd-1 that control programmed cell death.. We have identified and characterized a novel pro-apoptotic gene eif-3.K. Loss-of-function by mutation or RNAi inactivation in eif-3.K resulted in a decrease of cell corpses, whereas heatshock-induced over-expression of eif-3.K weakly but significantly increased cell corpses. Interestingly, the eif-3.K mutation partially suppressed ectopic cell deaths caused by over-expression of egl-1 or ced-4. This result suggests that eif-3.K may act downstream of or in parallel to egl-1 and ced-4 in the programmed cell death pathway. Using a cell-specific promoter to express eif-3.k in touch neurons, we showed that eif-3.K likely promoted cell death in a cell-autonomous manner. To further explore EIF-3.K function, we generated antibodies against bacterially expressed EIF-3.K protein. We found that EIF-3.K was ubiquitously expressed during embryogenesis and localized to the cytoplasm. As human eif-3.K can functionally substitute C. elegans eif-3.K in an eif-3.K mutant, the function of eif-3.K in apoptosis is likely conserved in evolution.

Chun-Yi Huang et al. (2006) East Asia C. elegans Meeting "eif-3.K is a pro-apoptotic gene in C. elegans"

Teng-Wei Huang, Chun-Yi Huang

[East Asia C. elegans Meeting, 2006]

Programmed cell death or apoptosis is an important feature during C. elegans development. The pro-apoptotic genes egl-1, ced-4 and ced-3 are required for the execution of cell death. We have identified and characterized a novel pro-apoptotic gene eif-3.K. A loss-of-function mutation or inactivation by RNA interference in eif-3.K resulted in a reduction of cell corpse number during embryogenesis, whereas heatshock-induced over-expression of eif-3.K weakly but significantly promoted programmed cell death. In addition, eif-3.K mutation partially suppressed ectopic cell deaths caused by over-expression of egl-1 and ced-4. This result suggests that eif-3.K may act downstream of or in parallel to egl-1 and ced-4 in the genetic pathway during programmed cell death. Using a cell-specific promoter we showed that eif-3.K likely promoted cell death in a cell-autonomous manner. We generated antibodies against bacterially expressed EIF-3.K protein. The immunostaining result showed that EIF-3.K was ubiquitously expressed during embryogenesis and localized to the cytoplasm. To better understand the cell-death defect of eif-3.K mutants, we are currently performing a 4D microscopic analysis of the cell death process in wild-type and eif-3.K mutants.

Huang CY et al. (2012) PLoS One "C. elegans EIF-3.K promotes programmed cell death through CED-3 caspase."

Tan CH, Huang CY, Wu SC, Chen JY, Wu YF, Wu YC, Chen RH, Lu PJ, Tzeng RY

[PLoS One, 2012]

Programmed cell death (apoptosis) is essential for the development and homeostasis of metazoans. The central step in the execution of programmed cell death is the activation of caspases. In C. elegans, the core cell death regulators EGL-1(a BH3 domain-containing protein), CED-9 (Bcl-2), and CED-4 (Apaf-1) act in an inhibitory cascade to activate the CED-3 caspase. Here we have identified an additional component eif-3.K (eukaryotic translation initiation factor 3 subunit k) that acts upstream of ced-3 to promote programmed cell death. The loss of eif-3.K reduced cell deaths in both somatic and germ cells, whereas the overexpression of eif-3.K resulted in a slight but significant increase in cell death. Using a cell-specific promoter, we show that eif-3.K promotes cell death in a cell-autonomous manner. In addition, the loss of eif-3.K significantly suppressed cell death-induced through the overexpression of ced-4, but not ced-3, indicating a distinct requirement for eif-3.K in apoptosis. Reciprocally, a loss of ced-3 suppressed cell death induced by the overexpression of eif-3.K. These results indicate that eif-3.K requires ced-3 to promote programmed cell death and that eif-3.K acts upstream of ced-3 to promote this process. The EIF-3.K protein is ubiquitously expressed in embryos and larvae and localizes to the cytoplasm. A structure-function analysis revealed that the 61 amino acid long WH domain of EIF-3.K, potentially involved in protein-DNA/RNA interactions, is both necessary and sufficient for the cell death-promoting activity of EIF-3.K. Because human eIF3k was able to partially substitute for C. elegans eif-3.K in the promotion of cell death, this WH domain-dependent EIF-3.K-mediated cell death process has potentially been conserved throughout evolution.

Wojtovich AP et al. (2011) PLoS One "SLO-2 is cytoprotective and contributes to mitochondrial potassium transport."

Sherman TA, Nehrke K, Urciuoli WR, Wojtovich AP, Nadtochiy SM, Brookes PS

[PLoS One, 2011]

Mitochondrial potassium channels are important mediators of cell protection against stress. The mitochondrial large-conductance "big" K(+) channel (mBK) mediates the evolutionarily-conserved process of anesthetic preconditioning (APC), wherein exposure to volatile anesthetics initiates protection against ischemic injury. Despite the role of the mBK in cardioprotection, the molecular identity of the channel remains unknown. We investigated the attributes of the mBK using C. elegans and mouse genetic models coupled with measurements of mitochondrial K(+) transport and APC. The canonical Ca(2+)-activated BK (or "maxi-K") channel SLO1 was dispensable for both mitochondrial K(+) transport and APC in both organisms. Instead, we found that the related but physiologically-distinct K(+) channel SLO2 was required, and that SLO2-dependent mitochondrial K(+) transport was triggered directly by volatile anesthetics. In addition, a SLO2 channel activator mimicked the protective effects of volatile anesthetics. These findings suggest that SLO2 contributes to protection from hypoxic injury by increasing the permeability of the mitochondrial inner membrane to K(+).

Johnson CK et al. (2020) J Neurophysiol "The Na+/K+-ATPase is needed in glia of touch receptors for responses to touch in C. elegans."

Wang Y, Bianchi L, Fernandez-Abascal J, Johnson CK, Wang L

[J Neurophysiol, 2020]

Four of the five types of mammalian mechanosensors are composed of nerve endings and accessory cells. In C. elegans we showed that glia supports the function of nose touch neurons via the activity of glial Na⁺ and K⁺channels. We show here that a third regulator of Na⁺ and K⁺, the Na⁺/K⁺-ATPase, is needed in glia of nose touch neurons for touch. Importantly, we show that the two Na⁺/K⁺-ATPase genes are needed for the function rather than structural integrity and that their ion transport activity is crucial for touch. Finally, when glial Na⁺/K⁺-ATPase genes are knocked-out, touch can be restored by activation of a third Na⁺/K⁺-ATPase. Taken together, these data show the requirement in glia of touch neurons of the function of the Na⁺/K⁺-ATPase. These data underscore the importance of the homeostasis of Na⁺ and K⁺, most likely in the space surrounding touch neurons, in touch sensation, a function that might be conserved across species.

Jospin M et al. (2002) J Physiol "Characterization of K(+) currents using an in situ patch clamp technique in body wall muscle cells from Caenorhabditis elegans."

Jospin M, Allard B, Segelat L, Mariol MC, Segalat L

[J Physiol, 2002]

The properties of K+ channels in body wall muscle cells acutely dissected from the nematode Caenorhabditis elegans were investigated at the macroscopic and unitary level using an in situ patch clamp technique. In the whole-cell configuration, depolarizations to potentials positive to -40 mV gave rise to outward currents resulting from the activation of two kinetically distinct voltage-dependent K+ currents: a fast activating and inactivating 4-aminopyridine-sensitive component and a slowly activating and maintained tetraethylammonium-sensitive component. In cell-attached patches, voltage-dependent K+ channels, with unitary conductances of 34 and 80 pS in the presence of 5 and 140 mm external K+, respectively, activated at membrane potentials positive to -40 mV. Excision revealed that these channels corresponded to Ca2+-activated K+ channels exhibiting an unusual sensitivity to internal Cl- and whose activity progressively decreased in inside-out conditions. After complete run-down of these channels, one third of inside-out patches displayed activity of another Ca2+-activated K+ channel of smaller unitary conductance (6 pS at 0 mV in the presence of 5 mm external K+). In providing a detailed description of native K+ currents in body wall muscle cells of C. elegans, this work lays the basis for further comparisons with mutants to assess the function of K+ channels in this model organism that is highly amenable to molecular and classical genetics.

Shibuya Y et al. (2022) Algorithms Mol Biol "Space-efficient representation of genomic k-mer count tables."

Kucherov G, Shibuya Y, Belazzougui D

[Algorithms Mol Biol, 2022]

MOTIVATION: k-mer counting is a common task in bioinformatic pipelines, with many dedicated tools available. Many of these tools produce in output k-mer count tables containing both k-mers and counts, easily reaching tens of GB. Furthermore, such tables do not support efficient random-access queries in general. RESULTS: In this work, we design an efficient representation of k-mer count tables supporting fast random-access queries. We propose to apply Compressed Static Functions (CSFs), with space proportional to the empirical zero-order entropy of the counts. For very skewed distributions, like those of k-mer counts in whole genomes, the only currently available implementation of CSFs does not provide a compact enough representation. By adding a Bloom filter to a CSF we obtain a Bloom-enhanced CSF (BCSF) effectively overcoming this limitation. Furthermore, by combining BCSFs with minimizer-based bucketing of k-mers, we build even smaller representations breaking the empirical entropy lower bound, for large enough k. We also extend these representations to the approximate case, gaining additional space. We experimentally validate these techniques on k-mer count tables of whole genomes (E. Coli and C. Elegans) and unassembled reads, as well as on k-mer document frequency tables for 29 E. Coli genomes. In the case of exact counts, our representation takes about a half of the space of the empirical entropy, for large enough k's.