Haslam SM et al. (2002) Biochem Soc Symp "The glycomes of Caenorhabditis elegans and other model organisms."

Haslam SM, Morris HR, Gems D, Dell A

[Biochem Soc Symp, 2002]

There is no doubt that the immense amount of information that is being generated by the initial sequencing and secondary interrogation of various genomes will change the face of glycobiological research. However, a major area of concern is that detailed structural knowledge of the ultimate products of genes that are identified as being involved in glycoconjugate biosynthesis is still limited. This is illustrated clearly by the nematode worm Caenorhabditis elegans, which was the first multicellular organism to have its entire genome sequenced. To date, only limited structural data on the glycosylated molecules of this organism have been reported. Our laboratory is addressing this problem by performing detailed MS structural characterization of the N-linked glycans of C. elegans; high-mannose structures dominate, with only minor amounts of complex-type structures. Novel, highly fucosylated truncated structures are also present which are difucosylated on the proximal N-acetylglucosamine of the chitobiose core as well as containing unusual Fuca1-2Gal1-2Man as peripheral structures. The implications of these results in terms of the identification of ligands for genomically predicted lectins and potential glycosyltransferases are discussed in this chapter. Current knowledge on the glycomes of other model organisms such as Dictyostelium discoideum, Saccaromyces cerevisiae and Drosophila melanogaster is also discussed

Haslam SM et al. (2003) Biochimie "Hallmarks of Caenorhabditis elegans N-glycosylation: complexity and controversy."

Dell A, Haslam SM

[Biochimie, 2003]

Caenorhabditis elegans has become one of the most widely used model organisms for a range of molecular cell biological applications and is being increasingly used by glycobiologists. However, a major problem has been the lack of knowledge of the structure of the protein-linked glycans from this organism. In recent years several groups have published structural data, particularly N-glycan structural data. However, some of these data are contradictory. In this review we critically assess all the N-glycan structural data and consider how close we are in our goal of defining the glycome of C. elegans.

Haslam SM et al. (1999) J Biol Chem "Structural studies of N-glycans of filarial parasites. Conservation of phosphorylcholine-substituted glycans among species and discovery of novel chito-oligomers."

Morris HR, Reason AJ, Dell A, Harnett W, Houston KM, Haslam SM

[J Biol Chem, 1999]

N-Type glycans containing phosphorylcholine (PC-glycans), unusual structures found in the important human pathogens filarial nematodes, represent a novel target for chemotherapy. Previous work in our laboratories produced compositional information on the PC-glycan of ES-62, a secreted protein of the rodent parasite Acanthocheilonema viteae. In particular, we established using fast atom bombardment mass spectrometry (MS) analysis that PC was attached to a glycan with a trimannosyl core, with and without core fucosylation, carrying between one and four additional N-acetylglucosamine residues. In the present study, we demonstrate that this structure is conserved among filarial nematodes, including the parasite of humans, Onchocerca volvulus, for which new drugs are most urgently sought. Furthermore, by employing a variety of procedures, including collision-activated dissociation MS-MS analysis and matrix-assisted laser desorption MS analysis, we reveal that surprisingly, filarial nematodes also contain N-linked glycans, the antennae of which are composed of chito-oligomers. To our knowledge, this is the first report of such structures in a eukaryotic glycoprotein.

Huitema K et al. (2004) EMBO J "Identification of a family of animal sphingomyelin synthases."

Huitema K, van den Dikkenberg J, Brouwers JF, Holthuis JC

[EMBO J, 2004]

Sphingomyelin (SM) is a major component of animal plasma membranes. Its production involves the transfer of phosphocholine from phosphatidylcholine onto ceramide, yielding diacylglycerol as a side product. This reaction is catalysed by SM synthase, an enzyme whose biological potential can be judged from the roles of diacylglycerol and ceramide as anti- and proapoptotic stimuli, respectively. SM synthesis occurs in the lumen of the Golgi as well as on the cell surface. As no gene for SM synthase has been cloned so far, it is unclear whether different enzymes are present at these locations. Using a functional cloning strategy in yeast, we identified a novel family of integral membrane proteins exhibiting all enzymatic features previously attributed to animal SM synthase. Strikingly, human, mouse and Caenorhabditis elegans genomes each contain at least two different SM synthase (SMS) genes. Whereas human SMS1 is localised to the Golgi, SMS2 resides primarily at the plasma membrane. Collectively, these findings open up important new avenues for studying sphingolipid function in animals.

Barbee SA et al. (2006) Dev Biol "The Sm proteins regulate germ cell specification during early C. elegans embryogenesis."

Evans TC, Barbee SA

[Dev Biol, 2006]

Sm and Sm-like proteins are core components of the splicesome but have other functions distinct from pre-mRNA processing. Here, we show that Sm proteins also regulate germ cell specification during early C. elegans embryogenesis. SmE and SmG were required to maintain transcriptional quiescence in embryonic germ cell precursors. In addition, depletion of SmE inhibited expression of the germ lineage-specific proteins PIE-1, GLD-1, and NOS-2, but did not affect maintenance of several maternal mRNAs. PIE-1 had previously been shown to activate transcriptional silencing and NOS-2 expression. We found that PIE-1 also promotes GLD-1 expression by a process that is independent of transcriptional silencing. Thus, Sm proteins could control transcriptional silencing and maternal protein expression by regulating PIE-1. However, loss of SmE function also caused defects in P granule localization and premature division in early germline blastomeres, processes that are independent of PIE-1 function. Therefore, the Sm proteins control multiple aspects of germ cell precursor development. Because depletion of several other core splicing factors did not affect these events, these Sm functions are likely distinct from pre-mRNA splicing. Sm family proteins assemble into ribonucleoprotein complexes (RNPs) that control RNA activities. We suggest that novel Sm RNPs directly or indirectly influence posttranscriptional control of maternal mRNAs to promote germ cell specification in the early C. elegans embryo.

Peg MacMorris et al. (2007) International Worm Meeting "A novel family of C. elegans snRNPs contains proteins associated with trans-splicing."

Madhur Kumar, Erika Lasda, Alison Larsen, Tom Blumenthal, Brian Kraemer, Peg MacMorris

[International Worm Meeting, 2007]

In approximately 70% of C. elegans pre-mRNAs, the RNA sequence between the 5 cap and the first 3 splice site is replaced by trans-splicing a short spliced leader (SL) from the Sm snRNP, SL1. C. elegans also utilizes a similar Sm snRNP, SL2, to trans-splice at sites between genes in polycistronic pre-mRNAs from operons. How do SL1 and SL2 snRNPs function in different contexts? Here we show that the SL1 snRNP contains a complex of SL75p and SL21p, homologs of novel proteins previously reported in the Ascaris SL snRNP. Interestingly, the SL2 snRNP does not contain either of these proteins. However, SL75p and SL26p, a paralog of SL21p, are components of another Sm snRNP that contains a novel snRNA species, Sm Y. Knockdown of SL75p is lethal. However, knockdown of either SL21p or SL26p alone leads to cold-sensitive sterility, whereas knockdown of both SL21p and SL26p is lethal. This suggests that these two proteins have overlapping functions even though they are associated with different classes of snRNP. These phenotypic relationships, along with the association of SL26p with SL75p imply that, like the SL1 RNA/Sm/SL75p/SL21p complex, the Sm Y/Sm/SL75p/SL26p complex is associated with trans-splicing. We hypothesize that the Sm Y snRNP is somehow associated with SL2-specific trans-splicing. This idea is supported by the fact that the sequences of SL2 RNA and Sm Y allow base pairing of the two snRNPs through their second stem/loops. We are currently testing for this base pairing interaction using psoralen/UV crosslinking.

Macmorris M et al. (2007) RNA "A novel family of C. elegans snRNPs contains proteins associated with trans-splicing."

MacMorris M, Larsen A, Lasda E, Blumenthal T, Kumar M, Kraemer B

[RNA, 2007]

In many Caenorhabditis elegans pre-mRNAs, the RNA sequence between the 5'' cap and the first 3'' splice site is replaced by trans-splicing a short spliced leader (SL) from the Sm snRNP, SL1. C. elegans also utilizes a similar Sm snRNP, SL2, to trans-splice at sites between genes in polycistronic pre-mRNAs from operons. How do SL1 and SL2 snRNPs function in different contexts? Here we show that the SL1 snRNP contains a complex of SL75p and SL21p, which are homologs of novel proteins previously reported in the Ascaris SL snRNP. Interestingly, we show that the SL2 snRNP does not contain these proteins. However, SL75p and SL26p, a paralog of SL21p, are components of another Sm snRNP that contains a novel snRNA species, Sm Y. Knockdown of SL75p is lethal. However, knockdown of either SL21p or SL26p alone leads to cold-sensitive sterility, whereas knockdown of both SL21p and SL26p is lethal. This suggests that these two proteins have overlapping functions even though they are associated with different classes of snRNP. These phenotypic relationships, along with the association of SL26p with SL75p, imply that, like the SL1 RNA/Sm/SL75p/SL21p complex, the Sm Y/Sm/SL75p/SL26p complex is associated with trans-splicing.

Scott A Barbee et al. (2001) International C. elegans Meeting "The splicesomal Sm proteins play a role in P granule localization during early embryogenesis."

Thomas C Evans, Scott A Barbee, Alex L Lublin

[International C. elegans Meeting, 2001]

In early C. elegans embryos, development is initiated by asymmetric cell division, which leads to the precise localization cell fate regulators to specific cells. One feature of early asymmetry is the localization of cytoplasmic RNP particles, the P granules, to the germ cell precursors at each of several polarized cell divisions. The composition and function of P-granules is not well understood. Further, the mechanisms that control the localization of P-granules and other factors in the embryo are poorly understood. We used RNA interference (RNAi) to screen a cDNA library for new genes involved in early polarity. Surprisingly, one of the genes identified is the ortholog of human SmE. SmE is one of several Sm proteins that form a multisubunit complex required for snRNP assembly, nuclear import, and mRNA splicing. RNAi of Sm subunits in C. elegans caused mislocalization of P granules to somatic sisters of germ cells after the 4-8 cell stages in the early embryo. In addition, Sm RNAi disrupted the subcellular distribution of PGL-1 (a P-granule component) in both mature germ cells and in germ cell precursors after the 8-cell stage. P-granules in wild type germ cells are primarily attached to the nucleus, but in Sm (RNAi) germ cells and embryos, the nuclear attachment of many P-granules is lost. At lower penetrance, GLP-1 protein was inappropriately expressed in posterior cells following Sm RNAi. By contrast, RNAi of the core splicing factors U1 70K and U2AF65, or of RNA polymerase II, had no effect on P granule segregation, subcellular distribution, or GLP-1 asymmetry, although all caused severe embryonic defects. These data suggest that the P-granule and GLP-1 asymmetry phenotypes from Sm RNAi are not likely to result from a general defect in splicing. Therefore, Sm proteins may have a role in P-granule localization that is independent of splicing. Interestingly, antibodies against the Sm complex stain P granules at all stages of development. Sm RNAi attenuates this staining. Therefore, Sm proteins may be P granule components that affect the localization of P granules by controlling their integrity or nuclear attachment in germ cells and their precursors. The Sm proteins are also found in the nucleus of most cells. However, nuclear localization of the Sm’s is dynamically regulated during oogenesis and germ cell precursor formation, suggesting that regulation of the Sm complex or snRNPs may be important for germ cell development.

Zhang W et al. (2016) Cell Mol Neurobiol "Shengmai Formula Ameliorates Pathological Characteristics in AD C.elegans."

Zhang W, Zhang Z, Wang X, Wang D, Zhi D, Ren H, Fei D, Zhu H, Li H

[Cell Mol Neurobiol, 2016]

Shengmai (SM) formula, a classical traditional Chinese medicine formula, is composed of Panax ginseng (Pg), Ophiopogon japonicus (Oj), and Schisandra Chinesis (Sc). SM has been clinically used to treat heart failure and ischemic heart disease. Although SM formula has been reported to be potential for fighting against Alzheimer's disease (AD) by previous works, there are many gaps in our knowledge on its usage in AD treatment on an organism level and will then need to be further clarified. In this study, transgenic Caenorhabditis elegans expressing human A1-42 are used to evaluate SM formula efficacy to treat AD phenotype and to investigate its underlying mechanism. The results showed that SM formula ameliorated AD pathological characteristics of paralysis behavior and chemotaxis defect in transgenic C.elegans. With SM treatment, the number of A deposits decreased, the levels of gene expressions of hsp16-2, hsp16-41, ace-1, ace-2, and TNFA1P1 homolog genes were down-regulated. Our results also showed that Oj exhibited more stronger effect on delaying paralysis in worms than Pg and Sc did, and synergistic action was observed between Pg and Oj, and Sc further enhanced the activity of Pg/Oj combination on delaying paralysis behavior. Further, SM with herbs of Pg, Oj, and Sc at a dose proportion of 9:9:6 exhibited superior therapeutic efficacy in comparison with herbs at other dose proportions. After SM formula extracted by ethanol, it delayed AD symptoms on a wider dose from 0.2 to 10.0mg/mL with no toxic effect. These results provided more evidence for SM formula being potential to be used to treat AD.

Scott A Barbee et al. (2003) International Worm Meeting "The spliceosomal Sm proteins regulate germ cell fate during early C. elegans embryogenesis."

Thomas C Evans, Scott A Barbee

[International Worm Meeting, 2003]

General mRNA processing factors are traditionally thought to function only in the control of global gene expression. However, we have previously shown that the Sm proteins, core components of the spliceosome, also regulate germ granule localization during early C. elegans embryogenesis independently of mRNA splicing. Here we provide evidence that the Sm proteins are involved in the specification of germ cells by regulating transcriptional quiescence and protein expression in germ cell precursors of the embryo. In early development of several metazoans, germ cell precursors are transcriptionally inactive. One indication of this general transcriptional repression is the absence of phosphorylation of the C-terminal domain of RNA polymerase II. When Sm protein activity was disrupted by RNA interference (RNAi), CTD phosphorylation occurred inappropriately in germ cell precursors. In addition, at least one germ cell promoter is inappropriately activated in germ cell precursors in SmE(RNAi) embryos. These results suggest that the Sm complex is necessary for transcriptional silencing in these cells. In early C. elegans development, inhibition of transcription in germ cell precursors requires PIE-1, a CCCH-type zinc-finger protein that control germ cell fate. Disruption of Sm protein activity causes reduction and possibly mislocalization of PIE-1 in early embryos. Loss of PIE-1 in SmE(RNAi) embryos occurs after the 1-cell stage, suggesting that translation or stability of maternal PIE-1 is regulated by the Sms in the embryo. Additionally, expression of NOS-2, a germ cell-specific factor whose translation requires PIE-1, is attenuated following Sm (RNAi). Taken together, this suggests that the Sm proteins may control germ cell fate by regulating PIE-1 expression. In contrast, loss of other splicing factors had little or no impact on each of the germ precursor markers. These observations suggest that the Sm proteins regulate germ cell fate or function in the early embryo by a mechanism distinct from their role in pre-mRNA splicing. Sm and Sm-related proteins bind small non-coding RNAs and possibly mRNAs. Therefore, the Sm complex may control germ cell fate and other processes as a novel broad-based chaperone for RNP assembly and/or RNA:RNA interactions.