Soderlund CA et al. (1990) Worm Breeder's Gazette "gm Sequence Analysis System Update"

Soderlund CA, Fields CA, Shanmugam P

[Worm Breeder's Gazette, 1990]

A number of improvements have been made in the gm automated DNA sequence analysis system: (1) The ratio of AT-containing to CG-containing dinucleotides has been added as a test for introns. This works better than AT frequency alone in C. elegans.(2) A branch site consensus sequence or an enhanced dinucleotide ratio can be required as an additional test on introns. (3) Predicted amino-acid sequence files are generated in a format appropriate for input to the Dana-Farber motif-identification program plsearch. (4) A graphic interface based on X-windows, version 11 is available as an option. (5) The complete analysis algorithm is significantly faster than the previous version. (6) A greedy model evaluation algorithm is available as an option. This algorithm generates the longest, non-overlapping models that cover a sequence and is much faster than the complete analysis algorithm. The program has been tested on Sun3, Sun4 and VAX machines running Unix (Ultrix on the VAX). Results for a series of tests run on a Sun 4/60 are shown in the table. [See Figure 1] gm can be run remotely on our machine, using the Internet. To do this, telnet to haywire.nmsu.edu, and logon as gm_guest with password gmuser. Read the README file for information on running gm. We are also distributing gm as C source code to nonprofit laboratories, either via anonymous ftp to haywire.nmsu.edu, or on tape. If you would like to receive gm on tape, send a 1/4' cartridge or 1/2' reel tape to: Chris Fields, Box 30001/3CRL, New Mexico State University, Las Cruces, New Mexico 88003-0001, USA; Telephone (505) 646-2848.

Soderlund CA et al. (1991) International C. elegans Meeting "FINE-TUNING gm, V. 2.0 FOR WORM SEQUENCE ANALYSIS."

Shanmugam P, Fields CA, Soderlund CA, White O

[International C. elegans Meeting, 1991]

As more worm genes are sequenced, considerable variability in the characteristics of both exons and introns at the DNA level is becoming apparent. Features of interest include: Length. Lengths of both exons and introns range from a few tens of bases to around 10 kb. Short exons are very common. While the peak of the intron size distribution is still at 50-52 nucleotides, introns with lengths greater than 1 kb are now fairly common. Base composition. Exons are richer in C and G than introns, but the average difference in composition between exons and introns is decreasing as more sequences become available. Long introns often have islands of relatively high C+G content. The dinucleotides AA and TT are, however, much more common than CC or GG in introns. Codon usage. Codon preferences differ significantly between gene farnilies. There is a general trend toward less asymmetrical codon preferences in weaklyexpressed gene families; however, there are also strong codon preference reservals in some gene families, e.g. glp-l - lin-12. Sites. S' splice-site sequences differ significantly between long and short introns. This variation, first observed in C. elegans, has now been noted in Drosophila and plants, and may be ubiquitous. 3' splice site sequences appear not to vary with intron length. Complexity. Regulatory genes appear to contain larger numbers of potential splice sites, and to have less-asymmetric base compositions between exons and introns, than highly-expressed structural-protein genes. The efficiency of correct splice-site selection may be affected by these features. The variability in these features significantly increases the difficulty of gene identification by sequence analysis. We have used C. elegans sequences extensively for testing our gm automated DNA sequence analysis software. A number of new functions have been incorporated into gm v. 2.0 to provide greater accuracy of predictions in the face of compositional and site variations between genes. These include more accurate site-identification methods that combine consensus matrix and compositional methods and flexible multinucleotide compositional measures. Functions have also been added that allow automated use of partial cDNA data for genomic sequence analysis. The performance of gm v. 2.0 is currently being evaluated on a large set of worm genes.

Wang QC et al. (2016) Cell "TMCO1 Is an ER Ca(2+) Load-Activated Ca(2+) Channel."

Li Y, Yang Y, Liu Y, Zhang B, Tang TS, Yu J, Tan H, Chen Q, Wang H, Yang M, Wang X, Zheng Q, Li X, Zhou A, Guo C, Wang F, Wang JQ, Chai H, Sun Z, Wang QC, Zhu S

[Cell, 2016]

Maintaining homeostasis of Ca(2+) stores in the endoplasmic reticulum (ER) is crucial for proper Ca(2+) signaling and key cellular functions. The Ca(2+)-release-activated Ca(2+) (CRAC) channel is responsible for Ca(2+) influx and refilling after store depletion, but how cells cope with excess Ca(2+) when ER stores are overloaded is unclear. We show that TMCO1 is an ER transmembrane protein that actively prevents Ca(2+) stores from overfilling, acting as what we term a "Ca(2+) load-activated Ca(2+) channel" or "CLAC" channel. TMCO1 undergoes reversible homotetramerization in response to ER Ca(2+) overloading anddisassembly upon Ca(2+) depletion and forms a Ca(2+)-selective ion channel on giant liposomes. TMCO1 knockout mice reproduce the main clinical features of human cerebrofaciothoracic (CFT) dysplasia spectrum, a developmental disorder linked to TMCO1 dysfunction, and exhibit severe mishandling of ER Ca(2+) in cells. Our findings indicate that TMCO1 provides a protective mechanism to prevent overfilling of ER stores with Ca(2+) ions.

Wang D et al. (2010) J Environ Sci (China) "Confirmation of combinational effects of calcium with other metals in a paper recycling mill effluent on nematode lifespan with toxicity identification evaluation method."

Wang D, Wang Y, Shen L

[J Environ Sci (China), 2010]

We used toxicity identification evaluation (TIE) method to confirm the combinational effects of identified toxic metals in a paper recycling mill effluent in inducing the decreased lifespan in nematode Caenorhabditis elegans. Exposure to Ca + Al caused more severely decreased lifespan than that exposed to Ca, or Al; and exposure to Ca + Fe induced more severely decreased lifespan than that exposed to Ca, or Fe. Exposure to Ca+Al+Fe caused more severely decreased lifespan than that exposed to Ca, or Ca+Fe. Moreover, the baseline toxicity on lifespan was doubled by doubling the concentration of combined metals (Ca+Al+Fe) in spiking test in original effluent (oe), and lifespan defects in oe+Ca+Al+Fe exposed nematodes were more severe than that in Ca+Al+Fe exposed nematode. Therefore, Ca+Al+Fe exposure may largely explain the formation of decreased lifespan induced by the examined industrial effluent. Furthermore, the observed reduction of lifespan induced by the combination of high level of Ca with other metals may be at least partially independent of the insulin-like pathway.

Narayanaswamy N et al. (2018) Nat Methods "A pH-correctable, DNA-based fluorescent reporter for organellar calcium."

Chakraborty K, Krishnan Y, Zeichner E, Devany J, Leung K, Narayanaswamy N, Saminathan A

[Nat Methods, 2018]

It is extremely challenging to quantitate lumenal Ca²⁺ in acidic Ca²⁺ stores of the cell because all Ca²⁺ indicators are pH sensitive, and Ca²⁺ transport is coupled to pH in acidic organelles. We have developed a fluorescent DNA-based reporter, CalipHluor, that is targetable to specific organelles. By ratiometrically reporting lumenal pH and Ca²⁺ simultaneously, CalipHluor functions as a pH-correctable Ca²⁺ reporter. By targeting CalipHluor to the endolysosomal pathway, we mapped lumenal Ca²⁺ changes during endosomal maturation and found a surge in lumenal Ca²⁺ specifically in lysosomes. Using lysosomal proteomics and genetic analysis, we found that catp-6, a Caenorhabditis elegans homolog of ATP13A2, was responsible for lysosomal Ca²⁺ accumulation-an example of a lysosome-specific Ca²⁺ importer in animals. By enabling the facile quantification of compartmentalized Ca²⁺, CalipHluor can expand the understanding of subcellular Ca²⁺ importers.

Takayama J et al. (2016) Cell Rep "The Sperm TRP-3 Channel Mediates the Onset of a Ca(2+) Wave in the Fertilized C.elegans Oocyte."

Takayama J, Onami S

[Cell Rep, 2016]

Sperm induce Ca(2+) waves in the fertilized egg by introducing soluble factors or by surface interactions, which activate egg Ca(2+) channels. Involvement of sperm Ca(2+) channels is predicted by the conduit model; however, this model has not been validated. In Caenorhabditis elegans, the sperm-specific TRP family Ca(2+) channel TRP-3 mediates sperm-oocyte fusion. Here, using high-speed invivo imaging and image analyses, we show that sperm induce an immediate local Ca(2+) rise followed by a Ca(2+) wave in fertilized C.elegans oocytes. Oocytes fertilized by rare trp-3 escaper sperm showed a lack of local rise and a delay in onset of the Ca(2+) wave. Sperm Ca(2+) imaging suggests that the local rise is not due to the bolus introduction of stored Ca(2+). These results suggest that, along with its primary function in sperm-oocyte fusion, TRP-3 induces Ca(2+) waves in fertilized oocytes, consistent with the conduit model.

Zajac M et al. (2024) Sci Adv "A mechanism of lysosomal calcium entry."

Saminathan A, Anees P, Mukherjee S, Srikumar J, Oettinger D, Henn K, Zajac M, Krishnan Y, Zou J

[Sci Adv, 2024]

Lysosomal calcium (Ca²⁺) release is critical to cell signaling and is mediated by well-known lysosomal Ca²⁺ channels. Yet, how lysosomes refill their Ca²⁺ remains hitherto undescribed. Here, from an RNA interference screen in <i>Caenorhabditis elegans</i>, we identify an evolutionarily conserved gene, <i>lci-1</i>, that facilitates lysosomal Ca²⁺ entry in <i>C. elegans</i> and mammalian cells. We found that its human homolog TMEM165, previously designated as a Ca²⁺/H⁺ exchanger, imports Ca²⁺ pH dependently into lysosomes. Using two-ion mapping and electrophysiology, we show that TMEM165, hereafter referred to as human LCI, acts as a proton-activated, lysosomal Ca²⁺ importer. Defects in lysosomal Ca²⁺ channels cause several neurodegenerative diseases, and knowledge of lysosomal Ca²⁺ importers may provide previously unidentified avenues to explore the physiology of Ca²⁺ channels.

Niwa F et al. (2016) Biochem Biophys Res Commun "Dissection of local Ca(2+) signals inside cytosol by ER-targeted Ca(2+) indicator."

Mikoshiba K, Kobayashi A, Niwa F, Sakuragi S, Bannai H, Oda Y, Takagi S

[Biochem Biophys Res Commun, 2016]

Calcium (Ca(2+)) is a versatile intracellular second messenger that operates in various signaling pathways leading to multiple biological outputs. The diversity of spatiotemporal patterns of Ca(2+) signals, generated by the coordination of Ca(2+) influx from the extracellular space and Ca(2+) release from the intracellular Ca(2+) store the endoplasmic reticulum (ER), is considered to underlie the diversity of biological outputs caused by a single signaling molecule. However, such Ca(2+) signaling diversity has not been well described because of technical limitations. Here, we describe a new method to report Ca(2+) signals at subcellular resolution. We report that OER-GCaMP6f, a genetically encoded Ca(2+) indicator (GECI) targeted to the outer ER membrane, can monitor Ca(2+) release from the ER at higher spatiotemporal resolution than conventional GCaMP6f. OER-GCaMP6f was used for in vivo Ca(2+) imaging of C. elegans. We also found that the spontaneous Ca(2+) elevation in cultured astrocytes reported by OER-GCaMP6f showed a distinct spatiotemporal pattern from that monitored by plasma membrane-targeted GCaMP6f (Lck-GCaMP6f); less frequent Ca(2+) signal was detected by OER-GCaMP6f, in spite of the fact that Ca(2+) release from the ER plays important roles in astrocytes. These findings suggest that targeting of GECIs to the ER outer membrane enables sensitive detection of Ca(2+) release from the ER at subcellular resolution, avoiding the diffusion of GECI and Ca(2+). Our results indicate that Ca(2+) imaging with OER-GCaMP6f in combination with Lck-GCaMP6f can contribute to describing the diversity of Ca(2+) signals, by enabling dissection of Ca(2+) signals at subcellular resolution.

Ronan, E.A. et al. (2019) International Worm Meeting "Neural and genetic mechanisms of cinnamaldehyde sensation in C. elegans."

Ronan, E.A., Guo, Y., Sartbaeva, E., Zhang, X., Liu, J., Xu, X.Z.S., He, F.

[International Worm Meeting, 2019]

Cinnamaldehyde (CA), the essential oil in cinnamon, is shown to have systemic metabolic benefits in both mice and humans, however the underlying molecular mechanisms are not understood. While the only known receptor for CA is transient receptor potential (TRP)A1, the thermogenic response of adipocytes to CA are TRPA1-independent, indicating unknown CA-sensing mechanisms exist. Here we report that CA acts on sensory neurons in C. elegans to evoke complex behavioral responses. In particular, we found that CA can excite the polymodal nociceptive neuron ASH shown by calcium imaging and electrophysiology recordings. We found these effects to be mediated by GPCR signaling, suggesting that the underlying CA receptor is a GPCR. We performed genetic screens to identify the genes mediating CA-avoidance in the ASH neuron. Our results show that C. elegans sense CA through a previously uncharacterized neural and genetic pathway which may be evolutionarily conserved and have physiological significance in higher organisms.

Estevez AY et al. (2005) J Physiol "Calcium feedback mechanisms regulate oscillatory activity of a TRP-like Ca2+ conductance in C. elegans intestinal cells."

Strange K, Estevez AY

[J Physiol, 2005]

Inositol-1,4,5-trisphosphate (IP3)-dependent Ca(2+) oscillations in Caenorhabditis elegans intestinal epithelial cells regulate the nematode defecation cycle. The role of plasma membrane ion channels in intestinal cell oscillatory Ca(2+) signaling is unknown. We have shown previously that cultured intestinal cells express a Ca(2+) selective conductance, IORCa, that is biophysically similar to TRPM7 currents. IORCa activates slowly and stabilizes when cells are patch clamped with pipette solutions containing 10 mM BAPTA and free Ca(2+) concentrations of ~17 nM. However, when BAPTA concentration is lowered to 1 mM, IORCa oscillates. Oscillations in channel activity induced simultaneous oscillations in cytoplasmic Ca(2+) levels. Removal of extracellular Ca(2+) inhibited IORCa oscillations whereas readdition of Ca(2+) to the bath caused a rapid and transient reactivation of the current. Experimental maneuvers that elevated intracellular Ca(2+) blocked current oscillations. Elevation of intracellular Ca(2+) in the presence of 10 mM BAPTA to block IORCa oscillations led to a dose-dependent increase in the rate of current activation. At intracellular Ca(2+) concentrations of 250 nM, current activation was transient. Patch pipette solutions buffered with 1-4 mM of either BAPTA or EGTA gave rise to similar patterns of IORCa oscillations. We conclude that changes in Ca(2+) concentration close to the intracellular opening of the channel pore regulate channel activity. Low concentrations of Ca(2+) activate ORCa. As Ca(2+) enters and accumulates near the pore mouth, channel activity is inhibited. Oscillating plasma membrane Ca(2+) entry may play a role in generating intracellular Ca(2+) oscillations that regulate the C. elegans defecation rhythm.