Friedlein A et al. (2000) Worm Breeder's Gazette "The Function of the Putative Cargo Receptor ERGIC-53 in the Worm"

Friedlein A, Hauri H-P, Lahm HW, Kuhn F

[Worm Breeder's Gazette, 2000]

Trafficking along the exocytic pathway involves a complex system of transport vesicles and membrane structures. The early secretory pathway includes the ER, the ER-Golgi intermediate compartment (ERGIC) and the Golgi apparatus that are connected by a vesicle-mediated anterograde (from the ER) and retrograde (to the ER) protein transport pathway. The ERGIC is now best defined by the marker protein ERGIC-53, a type I transmembrane protein.

Sigrist CB et al. (1999) Worm Breeder's Gazette "Vulva formation in Pristionchus pacificus relies on continuous gonadal induction."

Sommer RJ, Sigrist CB

[Worm Breeder's Gazette, 1999]

One of the best known features of vulva development in Caenorhabditis elegans is the induction of the vulval precursor cells by the gonadal anchor cell. Induction is crucial for the initiation of pattern formation within the C. elegans vulva equivalence group and therefore, it is surprising to find that this aspect of vulva formation in particular, varies greatly among nematodes. In some species which form vulvae in the posterior body region, no gonadal signal is necessary for vulva induction. In other nematodes, like Panagrolaimus, Oscheius and Rhabditella, vulva formation depends on two temporally distinct gonadal inductions which specify the different cell fates.

Dent JA et al. (1998) Worm Breeder's Gazette "Ivermectin as a Paralytic Agent"

Dent JA, Avery L

[Worm Breeder's Gazette, 1998]

We have found it difficult to paralyze worms for the purpose of visualizing and/or photographing GFP expression. The best agent for paralyzing worms, sodium azide, quenches GFP fluorescence. We have tried 1-phenoxy-2-propanol, ethanol and levamisol and have found that effective doses of these drugs significantly distort the worm's morphology. Ivermectin is currently our paralytic of choice. Worms soaked in M9 with 1% DMSO and 100 ng/ml ivermectin for a couple hours are generally inert without significant morphological distortion apart from the occasional vacuole. The worms can also be treated by placing them on ivermectin plates (containing 1% DMSO and 100ng/ml ivermectin) which takes longer but allows you to look at them to see when they become paralyzed. Eggs of paralyzed worms can always be recovered and sometimes adults, when not too far gone, can recover from ivermectin.

Fodor A (1980) Worm Breeder's Gazette "Letter Addressed to B. Edgar Regarding New Data on Bal-X-1"

Fodor A

[Worm Breeder's Gazette, 1980]

Dear Dr. Edgar: 28.11.1979. Probably you have got our two papers for News Letters (dealing with Precocene in Nematodes and Bal-X-chromosome, respectively. I should like to complete the Bal-X-1 paper with some recent data arriving from Szeged today. The genotype of hermaphrodites of the stock: dpy-8( e1321) * unc-3(e151) over * lon-2(e678) *.The distribution of the self- progeny was the followings: [See Figure 1] The occurrence of males having one of the X chromosomes alternatively among P1 progeny indicate the the presence of Bal-X-1 chromosome causes some anomaly in meiotic pairing of the homologs.The reason of this could be i.) The presence of a dominant him mutation ( homozygous lethal) nearby to the lon 2 marker; ii.) chromosome aberration, probably translocation. Considering,that we could not manage to ride of this dominant effect by several outcrosses ( recombination) we prefer the second hypothesis to test. Best regards: Andras Fodor (BRC, Szeged)

Gibert M-A et al. (1978) Worm Breeder's Gazette "technical note on: use of a modified M9 buffer as an incubation medium for C elegans ovotestis."

Gibert M-A, Starck J

[Worm Breeder's Gazette, 1978]

For radioautographic studies of in vitro RNA synthesis during C. elegans oogenesis, J. Starck used M9 buffer (BRENNER, 1974) as an incubation medium. The results of labelling were rather good, they showed a mode of RNA synthesis which should correspond to a normal metabolism, (J. Starck, 1977). The longer incubation time was two hours. Electron microscopic studies showed that subcellular structures were not keep good we often remarked broken nuclear membranes and mitochondria as swelled nuclei. The oocytes seemed to be hypertrophied and parietal cells were crushed. These observations became more numerous when the incubation times became longer. Furthermore, systematic studies on fixatives showed that the best one has an osmotic pression around 400 mOsM the M9 buffer osmotic pression is only about 300 mOsM. We try to make better the incubation medium by adding saccharose. With 3% of this product, the osmotic pression of the medium, called M10, is about 400 mOsM. New essays with this M10 gave the same results in photonic radioautographies. In electron microscopic studies, we note a normal cellular ultrastructure, without accidents on nuclei or mitochondria.

Starck J et al. (1980) Worm Breeder's Gazette "analysis of proteins from larvae and adults of C elegans."

Starck J, Bosch C

[Worm Breeder's Gazette, 1980]

Extracts from larvae and adults were prepared using different methods: Animals were resuspended in buffer and homogenized either with a dounce, in a mortar with an equal weight of powdered glass or by sonication. The fourth method involved direct treatment of the animals for 10 min at 100 C in the presence of SDS and - mercaptoethanol. After centrifugation to remove large debris, the suspension (with addition of supplementary -mercaptoethanol in the last method) was analyzed on a polyacrylamide gradient (3.75 to 15 %) in the presence of urea and SDS. After electrophoresis, proteins were stained with Coomassie blue. We observed some variations in the protein pattern depending on the technique used for solubilization. Patterns allowing comparison of adults and larvae were best obtained when sonication was employed to disrupt the animals. Under such conditions, two high molecular weights (> 100,000 daltons) bands are present in adults but never in larvae. These bands look similar to the one found by Klass et al. (Dev. Biol., 1979, 69, 329-335) using labelled polypeptides. Caracterization of these two polypeptides is in progress.

Rosenquist TA et al. (1987) Worm Breeder's Gazette "a lucky artifact."

Kimble JE, Rosenquist TA

[Worm Breeder's Gazette, 1987]

We have inadvertently isolated a partial cDNA clone to a C. elegans protein kinase. The amino acid sequence deduced from the nucleotide sequence of this clone is shown below aligned with several protein kinases. The clone contains two sequence elements (DFG and APE) that are diagnostic of protein kinases. A third diagnostic element (RDL) would not lie within the peptide encoded by this clone. These three sequence elements are believed to fall within the active site of protein kinases. This clone lacks a tyrosine residue about 14 amino acids upstream of the APE box which is a hallmark of protein tyrosine kinases. In addition, Protein Kinase C gives the best alignment scores with this sequence. These observations lead us to believe that this clone is from a serine or threonine kinase. The sequence of a larger clone is needed to identify the specific kinase class to which this protein belongs. If anybody wishes to collaborate with us in the characterization of this protein kinase please give us a call. [See Figure 1]

Lewis JA (1978) Worm Breeder's Gazette "a convenient way of preparing radioactive worms."

Lewis JA

[Worm Breeder's Gazette, 1978]

We have tried a number of different ways of growing radioactive worms including forced aeration, rotary shaking and growth on Petri plates made with radioactive sulfate. The method we like best and which seems to give a good yield of healthy worms (mutant as well as wild-type) is the following: We grow [35S]-labelled bacteria (0.5 mCi/100 ml of 0.05 mM sulfate) by rotary shaking, collect the bacteria by centrifugation, resuspend them in 15 ml of S medium and divide the concentrated bacteria between two large empty Petri plates. A chunk of worm-filled agar is added and the plates taped shut with masking tape (no apparent anoxia for up to two weeks-incomplete taping is used to prevent desiccation of stock plates at 15 C). This method is convenient for doing a large number of preparations simultaneously at controlled temperatures and the worms are in a reasonably small volume for harvesting. To maximize incorporation, it is best to gauge the amount of bacteria left by carefully tilting a plate under a dissecting microscope (precocious plates are kept at 4 C up to 24 hours). To obtain dauer larvae, the plates are harvested two weeks after innoculation. We have a modified method of harvesting worms. Flotation on 35% sucrose results in a substantial loss of worms ( especially young larvae) by osmotic shock. Flotation on Ficoli 400 of equivalent density does not work, either for reasons of viscosity or lack of osmotic shock to the bacteria. We collect our worms by spinning them at 300 x g for 5 minutes (1000 rmp in a Sorvall SS-34 head). The worms are then layered on a 14.8% w/w Ficoll 400 and sedimented for 15 minutes at 300 x g. Bacteria, worm cuticles and almost all dead and grotty alive old worms remain on top. Healthy larvae, adults and bunches of eggs are pelleted. Fungal contamination is not separable from worms by this method. If the worms are allowed to starve very much, adults may fail to sediment (not carefully examined). Dauer larvae are harvested by initial collection at 300 x g, as above, but purified by flotation for 5 minutes on 30% w/v sucrose at 1000 x g. The dauer larvae so obtained are essentially free of other worm stages and of contamination. Dauer larvae are useful to us because they are a relatively defined juvenile stage and seem to give more reproducible electrophoretic patterns of detergent-extractable proteins than mixed populations do. The patterns of detergent- extractable proteins from dauer larvae and well-fed worms show many quantitative and qualitative differences.

Coulson AR et al. (1989) Worm Breeder's Gazette "genome map."

Ameer H, Albertson DG, Sulston JE, Coulson AR, Waterston RH, Fishpool RM

[Worm Breeder's Gazette, 1989]

The map is now widely distributed electronically (see WBG 10(3), 67), but we are once again providing a summary for the gazette in the form of an output from the routine CHPLT. Do note that this is a provisional best guess, and that some linkages may later go away: please enquire if you need to know about the status of particular areas. When you receive cosmid clones, as stabs, please IMMEDIATELY streak them out on selective medium, pick small colonies, and grow 4ml minipreps (protocol from Alan Coulson if needed). For some cosmids, larger preps are liable to yield deleted DNA. Check that cosmid DNA appears full size (runs slower than lambda on agarose gels), then freeze a sample of good cells in 20% glycerol at -70 C. MRC computer account 'ARC' does not exist; Alan and John share account JES. A database node is now open at Seattle: modem number 206-467-2957; operator Phil Meneely. The summary of clone types given on the next page may be helpful when you are deciding which clones to request for your research. To reveal the most suitable clones for microinjection, the buried clones need to be displayed by the routine CONTASS; we will help you to do this if you ask. [See Figures 1- 3]

Tarsitano M et al. (2000) Worm Breeder's Gazette "Mammalian Growth Factors in C. elegans (and vice versa)"

McGhee JD, Persico G, Fukushige T, Signore M, Mangogna M, Minchiotti G, Bazzicalupo P, Tarsitano M

[Worm Breeder's Gazette, 2000]

PLGF (Placenta derived growth factor) is a member of a family of structurally related angiogenic growth factors, all containing the 8 cysteine residues of the Platelet derived growth factor (PDGF) motif. The family includes VEGF, VEGF-B,VEGF-C and VEGF-D. Knock-out mice are viable but show subtle vascular phenotypes as adults (Carmeliet and Persico, unpublished). Plgf also has chemotactic activity on non-endothelial cells. A search of the C. elegans genomic sequence turned up one possible candidate homolog on YAC Y119D3.663; a number of cDNAs had been identified and the longest of such cDNA clones (yk305g9) was kindly provided by Yuji Kohara. The 297-aminoacid sequence derived from the full size cDNA differs from the Genefinder prediction and shows 33% identity and 46% simlarity with mouse VEGF-C, 24% identity and 40% similarity with mouse VEGF-B and 27% identity and 47% simlarity with Xenopus PDGF-B. Alignments centre on the conserved cysteine residues and are shown below. Note that the spacing of the 8 cysteine residues are the main feature conserved even between closely related mammalian species (1). A reasonable alignment can be continued C-terminal to the conserved 8-cysteine region and suggests that this particular C. elegans molecule can be aligned best with members of the family that bind to heparin.