Jones AK et al. (1995) Worm Breeder's Gazette "Can bli-4 Complement S. cerevisiae kex2 Mutants?"

Jones AK, Boone C, Thacker CM, Rose AM

[Worm Breeder's Gazette, 1995]

Can bli-4 Complement S.cerevisiae kex2 Mutants?

Harfe BD et al. (1998) Worm Breeder's Gazette "Neuronal specific bsh homeobox gene needs a home, inquire within."

Fire A, Harfe BD

[Worm Breeder's Gazette, 1998]

Over the last several years we have been characterizing the NK-2 homeodomain family in C. elegans. During the course of this work we have also studied a related homeobox gene, since named ceh-29. CEH-29 is a member of the bsh (brain specific homeobox) family (thanks to T. Burglin for help in classification). The founding member of this family, Drosophila bsh, is expressed exclusively in the brain where it is thought to play a highly specialized role in the determination and function of brain cell types (Jones and McGinnis, 1993). ceh-29 was initially identified by the genome sequencing project. The gene maps to chromosome II (cosmid F31E8) between dpy-10 and cyp-10 with the gene snt-1 located ~7kb downstream. As a starting point in our analysis of ceh-29 we used phage libraries to isolate genomic DNA and a putative full length cDNA. The cDNA clone allowed us to determine that CEH-29 was a relatively small protein (190aa), containing a homeodomain and a number of acidic regions. To determine where the ceh-29 promoter was active, we created ceh-29::gfp and lacZ fusions. In transgenic lines, reporter expression was first detected during early embryogenesis (~100 cells) in what appeared to be neuronal precursors. Reporter expression continued in neuronal cells reaching very high levels. In early larval development, cell proccesses of numerous neural cells could be clearly observed. To determine the consequences of loss of function of ceh-29, we carried out ceh-29 RNAi in wild-type animals. The F1 offspring of injected animals were then scored for visible phenotypes. In these animals we observed a low penetrance L1 scrawny arrest phenotype, with the majority appearing wild-type We have not characterized this gene further. Anyone interested in analyzing the neuronal ceh-29::gfp expression pattern, or further investigating the ceh-29 loss of function phenotype should contact us. B. Jones and W. McGinnis (1993). A new Drosophila homeobox gene, bsh, is expressed in a subset of brain cells during embryogenesis. Development 117, 793-806.

Gharib S et al. (2000) Worm Breeder's Gazette "Correlating the C. elegans and C. briggsae genetic maps"

Gharib S, Ally A, Baillie DL, Sternberg PW, Johnsen B, Brown KB

[Worm Breeder's Gazette, 2000]

We are developing a genetic map of Caenorhabditis briggsae in order to facilitate comparative studies with Caenorhabditis elegans. The two nematode species are morphologically similar but are approximately 25 - 40 million years apart. The comparison of the genetic maps of the two species should reveal how selective pressure constrains evolution of the linkage groups. For example, in C. elegans chromosomes there are regions of low and high rates of recombination (R. Johnsen Ph.D Thesis 1990, Simon Fraser University, Burnaby, Canada; S. Jones Ph.D. Thesis 1999 Cambridge Eng.). It has been postulated that highly conserved genes are more likely to be in regions of low recombination whereas rapidly evolving genes more likely reside in regions of high recombination. If this were true we would expect a similar gene distribution in C. briggsae. Our studies should also prove valuable for researchers who are comparing specific genes in in two nemotode species. As part of our investigations we want to identify which C. elegans genes correspond to each of the C. briggsae genes. So far we have identified 15 cby, 7 mip, 1 rot and 1 sml genes (the C. briggsae versions of dpy, unc, rol and sma respectively). Five of the cby genes, three of the mips and the rot are on the X-chromosome while the other 10 cbys, 4 mips and the sml gene map to autosomes. We have concluded that mip-1 is the C. briggsae version of unc-22 and cby-4 is dpy-1. We have also shown that mip-1, mip-3, mip-6, cby-7 and cby-8 are linked. In order to correlate the two genetic maps in an efficacious manner we request any help that the members of the worm community can afford us. Either by way of rescuing DNA or by helping with the identification of phenotypes that requires special knowledge or conditions to identify.

Consortium TCeGS (1995) Worm Breeder's Gazette "The C. elegans Genome Sequencing Project: A Progress Report."

Consortium TCeGS

[Worm Breeder's Gazette, 1995]

The C. elegans genome sequencing project: A progress report. The C. elegans Genome Consortium Genome Sequencing Center, Washington University School of Medicine, St. Louis, Missouri, USA and Sanger Centre, Hinxton Hall, Cambridge, UK. We present here another progress report for the C. elegans genome sequencing project. The majority of chromosome I*II is now essentially complete, although some gaps remain where cosmids were not available (Figure 1). While we are rescuing these regions from YACs and lambda clones, we will continue to sequence the cosmid-rich regions of other chromosomes. In the previous issue of the WBG, we reported significant protein similarities (blastx score >100) for several cosmids in the central region of chromosome II. Since then, we have made considerable progress on much of the remainder of this region. Protein similarities for several additional cosmids from chromosome II are presented in Table 1 (all cosmids for which shotgun sequencing has been completed). We also have begun sequencing cosmids near the center of the X chromosome. The St. Louis group started with cosmid C23F12 (near kin-11) and are moving right to left along the physical map. The Sanger Centre group are moving left to right from the same starting point. Similarity data for X chromosome cosmids which have been processed through the initial shotgun phase are also included in Table 1. For each putative genomic locus, the highest blastx score and a brief identifier are listed. Although the cosmids which contain database hits may not be completely sequenced, the Consortium will make preliminary sequence data available to the community with the caveat that it is preliminary and may still contain errors. Finished cosmid sequences are now available by anonymous ftp at: ftp.sanger.ac.uk (directory: pub/C.elegans_sequences). For further information on blastx similarities, please contact LaDeana Hillier (lhillier@watson.wustl.edu) or Steve Jones (sjj@sanger.ac.uk). For information on sequencing plans or estimated completion times, please contact Richard Wilson (rwilson@watson.wustl.edu) or Alan Coulson (alan@sanger.ac.uk). All requests for cosmid clones should be addressed to Alan Coulson. For further information about the C. elegans genome project including our policy statement about sharing both data and sequencing expertise, please contact Richard Wilson.

Fabio Piano et al. (2001) Worm Breeder's Gazette "On the reproducibility of large-scale - RNAi screens"

Aaron Schetter, Fabio Piano, Ken Kemphues, Kris Gunsalus, Valerie Reinke, Stuart Kim, Diane Morton

[Worm Breeder's Gazette, 2001]

RNAi is being used routinely to determine loss-of-function phenotypes and recently large-scale RNAi analyses have been reported (1,2,3). Although there is no question about the value of this approach in functional genomics, there has been little opportunity to evaluate reproducibility of these results. We are engaged in RNAi analysis of a set of 762 genes that are differentially expressed in the germline as compared to the soma (4 -- "Germline"), and have reached a point in our analysis that allows us to look at the issue of reproducibility. We have compared the RNAi results of genes in our set that were also analyzed by either Fraser et al. (1 -- Chromosome 1 set "C1") or Gonczy et al. (2 -- Chromosome 3 set "C3"). In making the comparison we have taken into account the different operational definition of "embryonic lethal" used by the three groups. In the C3 study, lethal was scored only if there were fewer than 10 surviving larva on the test plate, or roughly 90% lethal. In our screen and the C1 screen the percent survival was determined for each test. To minimize the contribution of false positives from our set, in our comparison with the C1 set we defined our genes as "embryonic lethal" if at least 30% of the embryos did not hatch, but included all lethals defined by Fraser et al. (> 10%). For our comparison with the C3 set, we used a more restrictive definition of "embryonic lethal" that required that 90% of the embryos did not hatch. (This means that in Table 1, five genes from our screen that gave lethality between 30-90% were included in the not lethal category; one of these was scored as lethal by Gonczy et al.). We have analyzed 149 genes from the germline set that overlap with the C1 set and 132 genes that overlap with the C3 set. The table below shows the number of genes scored as embryonic lethal (EL) or not embryonic lethal (NL) in each study. (Note that these comparisons do not include data from our published collection of ovary-expressed cDNAs.) Table 1. Comparing RNAi analysis of the same genes in different studies. Germline Chromosome 1 Germline Chromosome 3 NL (117) EL (32) NL (97) EL (35) NL (104) 100 4 NL (89) 87 2 EL (45) 17 28 EL (43) 10 33 Overall, the degree of reproducibility is high. The concordance between our results and the published results was 86% with C1 (128/149 genes) and 90% with C3 (120/132). However, we scored a larger number of genes as giving rise to embryonic lethal phenotypes than the other studies did. What does this mean? One possibility is that we are generating a large number of false positives (God forbid!). The other interpretation is that there is a fairly high frequency of false negatives in each screen (4-8% in our screen (2/45; 4/49); 22% in the C3 screen (10/45); and 35% (17/49) in the C1 screen). It is no surprise that the different methods used by the three groups resulted in slightly different outcomes and we can only speculate on which methodological variation contributed most. In comparing our methods to those used in the C3 study we note that our two groups used different primer pairs for each gene; that we tested genes individually while they tested genes in pairs; and that the operational definition of "embryonic lethal" differed. Considering the latter two differences, we speculate that even with pools of two, the competition noted by Gonczy et al. in dsRNA pools could reduce levels of lethality below the 90% cutoff. The major difference between our approach and the C1 approach is feeding vs. injection, raising the possibility that for some genes feeding may be a less effective means of administering dsRNA. Whatever the basis for the difference, these comparisons indicate that genes scored as "non-lethal" in any single study may show an embryonic lethal RNAi phenotype when reanalyzed. It therefore seems useful to have more than one pass at analyzing C. elegans genes via RNAi. We are indebted to P. Gonczy for very useful comments. Fraser, A. G., Kamath, R. S., Zipperlen, P., Martinez-Campos, M., Sohrmann, M. and Ahringer, J. (2000). Functional genomic analysis of C. elegans chromosome I by systematic RNA interference. Nature 408 , 325-330. Gonczy, P., Echeverri, G., Oegema, K., Coulson, A., Jones, S. J., Copley, R. R., Duperon, J., Oegema, J., Brehm, M., Cassin, E. et al. (2000). Functional genomic analysis of cell division in C. elegans using RNAi of genes on chromosome III. Nature 408 , 331-336. Piano, F., Schetter, A. J., Mangone, M., Stein, L. and Kemphues, K. J. (2000). RNAi analysis of genes expressed in the ovary of Caenorhabditis elegans. Curr Biol 10 , 1619-1622. Reinke, V., Smith, H. E., Nance, J., Wang, J., Van Doren, C., Begley, R., Jones, S. J., Davis, E. B., Scherer, S., Ward, S. et al. (2000). A global profile of germline gene expression in C. elegans. Mol Cell 6 , 605-616.

Buechner M et al. (1997) Worm Breeder's Gazette "exc Gene Interactions"

Buechner M, Hedgecock EM

[Worm Breeder's Gazette, 1997]

We have continued genetic studies on exc mutants, in which the long hollow processes (canals) of the excretory cell form large fluid-filled cysts. We previously classified these mutants into five groups based on cyst morphology: exc-2, exc-4, let-4, and let-653 mutants form large septate cysts near the excretory cell body; exc-1 and exc-5 mutants form extremely large septate cysts (often visible via dissecting microscope) predominantly at the shortened canal termini; exc-3, exc-6, and exc-8 mutants show regional canal widenings and meanderings, and occasional inappropriate extra branchings to form multiple canal lumena; alleles of sma-1 exhibit a short, very wide, unseptate canal. Finally, the lone exc-7 allele rh252 exhibits small cysts along the length of the canal, terminating in a large group of tiny cysts. We have now made double mutants of several of the genes from various groups. Double mutants made of genes from within any one group showed no increase in severity in canal phenotype. For example, exc-2 (rh90) exc-4 (rh133) animals showed no changes in cyst size or morphology compared to rh90 or rh133 alone, and no increase in larval lethality. Similarly, exc-6 (rh103) exc-8 (rh210) canals appeared no shorter or more meandering than those of the single mutants, and exc-1 (rh26) exc-5 (rh232) mutant canals had similar cyst size and placement as the single mutants alone. Doubles constructed with exc-7 (rh252) showed interesting effects, however. exc-7 is epistatic to exc-6; the double mutants showed no abnormal branchings, only a series of small cysts along the entire canal length. Perhaps not surprisingly, the large cysts of exc-2 and exc-4 are epistatic to exc-7. Most strikingly, the doubles exc-7 (rh252) sma-1 (e30) and especially exc-7 (rh252) exc-3 (rh207) show canal phenotypes different from that of the single mutants; instead, these animals show large cysts near the excretory cell body, like those of exc-4 mutants. Finally, we have been unable so far to isolate doubles of exc-7 (rh252) and either exc-1 (rh26) or exc-5 (rh232); this could indicate that these mutants are lethal in combination. Our previous ultrastructural data, along with sequencing results from the labs of David Baillie (Jones & Baillie, Mol. Gen. Genet. 248, 719-726, !95) and Judith Austin (see McKeown, Patel, Praitis, and Austin, WBG 14 (2), p. 83) suggest that the exc genes encode proteins necessary for the proper placement of structural elements, both extracellular and cytoskeletal, at the apical epithelial surface. We believe that these new genetic results indicate that the differing morphologies of the various groupings of exc genes reflect different elements that interact with each other to shape this surface; Exc-7 may play a central position in these interactions.

Bird DM et al. (1991) Worm Breeder's Gazette "A Model for the Structure of the dpy-13 Collagen"

Bird DM, Chen TY

[Worm Breeder's Gazette, 1991]

Using a combination of computer and hand alignments, we have generated a model for the dpy-13 product in which the peptide folds back on itself to form a triple helical structure with non-helical termini (telopeptides) and a region of potential interaction with other collagens. This model is consistent with a proposal based on a classical, physical biochemical analysis of Ascaris collagen (1) ( McBride and Harrington, Biochem., 6: 1484, 1967). Key features of our model are: 1) The four stretches of sequence able to form triple-helix define three domains (3, 5 and 7 in Fig. 1). Domains 5 and 7 are available to participate in helix formation along most of their length, but domain 3 is sufficient to interact with only the amino-half of 5 and the carboxy half of 7. Perhaps the hypervariable domain (2) interacts with the remainder of domains 5 and 7. Alternatively, another peptide (8) with a gly-X-Y motif might contribute to triple-helix. The discontinuity in 5 possibly is required for such interactions. This folding arrangement places the second gly-X-Y region (5) in anti-parallel to the other two (3 and 7), effectively reversing X and Y order. This reversal enhances the preferential distribution of the bulky residues in the X position (Traub, Israel J. Chem., 12: 435, 1974) and places six of the seven proline residues in suitable locations for X/Y pairing (Jones and Miller, JMB, 218: 209, 1991). 2) Rather than being formed from the termini of peptides, the telopeptides are generated by turns in the non-gly-X-Y regions (4, 6 in Fig. l) and stabilized by disulfide linkages; the location of cysteines is consistent with formation of disulfide bridges between the ends of 1 and 5, and start of 5 and the end of 7. Chou/Fasman algorithms predict alpha-helix at both turns. The distribution of lysine residues in these turns is characteristic of other telopeptides. . 3) Sequences towards the amino terminus are not shown as being involved in folding as they presumably include a signal sequence and pro-collagen domain that are absent in the mature protein. It is not inconceivable that the putative pro-collagen peptidase target is the 'Box A consensus' (Levy, et al., WBG 11(5): 36) in the center of the otherwise hypervariable region 2. 'Box A' also contains a putative glycosylation site. It has been suggested that the semi-dominant alleles of dpy-13 (e.g. e184) might have products with aberrant collagen (triple helix?) formation but normal sites of interaction with other cuticle components, whereas the products of recessive alleles (e.g. e225) might have normal collagen helix but an altered ability to interact with other components (von Mende and Riddle,WBG 11(4): 50). The molecular location of these defects is consistent with our folding model. 1. The size of collagen monomers calculated by these workers is not in accord with that predicted from C. elegans gene sequence data. We feel that the conditions used for their sedimentation studies (low concentrations of reducing agents) were conducive to cross-linking of monomers. [See Figure 1]

Massie MR et al. (1997) Worm Breeder's Gazette "Exposure To Sodium Azide Induces Thermotolerance"

White GE, Stine KE, Reagan A, Boggs KD, Massie MR, Gall S

[Worm Breeder's Gazette, 1997]

Induction of heat shock proteins in response to thermal stress is a well-characterized phenomenon reported to occur in many organisms including C. elegans (Heschl and Baillie, DNA 8: 233-43, 1989). Other stressors, including chemical stressors, have also been shown to induce these proteins (D. Jones et al., Toxicology 109: 119-27, 1996). We have hypothesized that chemicals affecting energy metabolism will also induce the heat shock response. As a test compound, we chose sodium azide, an inhibitor of oxidative phosphorylation, that is also used as a nematode anesthetic. We have found that exposure to low levels of this chemical confers thermotolerance in the nematode. To test our hypothesis, we developed the following protocol: the wild type (N2) strain received a 1 hr pre-treatment with either azide or 33oC. After a 4 hr recovery at 22oC, nematodes were heat shocked at 37oC for 2 hrs. After heat shock, worms recovered again at 22oC for 4 hrs before the number of surviving worms was determined. A 4 hr recovery period was necessary because worms are not moving immediately after the 37oC heat shock. Additionally, we have observed that temperature fluctuation as little as 1.5oC can critically affect the outcome. For example, temperatures rising to 38.5oC for 4 minutes prove lethal to all worms. Azide concentrations of 5.0-20mM were chosen based on preliminary studies which indicated that concentrations >20mM were lethal. The control worms (receiving only the heat shock) had a 0.08 survival probability while animals receiving the 33oC pre-treatment had a 0.89 survival probability, illustrating the classic heat shock response. At all concentrations, the sodium azide pre-treated worms showed an increased survival probability when compared to the control. 10mM (0.07%) sodium azide, corresponding to the concentration typically used as an anesthetic, conferred maximal (0.70) survival probability. We are currently using Western Blots to test our hypothesis that the molecular mechanism of the response to azide is the same as the response to elevated temperatures. The hsps we are examining include hsp90, 70, 60, and 16 (hsp16 antibody kindly provided by Peter Candido). We have also been searching for hsp mutants. Elizabeth Malone and Jim Thomas reported at the worm meeting that daf-21 is really a hsp90 mutation. We are currently testing their temperature sensitive mutant (JT6130). Grown at the permissive temperature of 16oC, JT6130 survived heat shock after receiving pre-treatment with either azide or 33oC. However, these worms showed a reduction in survival probability when compared to N2. At the restricted temperature of 25oC, the worms pre-treated with either azide or 33oC did not survive the heat shock. These results strongly suggest that hsp90 is an important part of the worm's response to stress. We are searching ACEDB, as well as the literature, for information on mutants with defects in some aspect of energy metabolism. Like daf-21 and hsp90, we suspect that mutations in energy metabolism proteins may have already been identified by other names; therefore, we welcome any additional information concerning mutants that are thought to be involved in energy metabolism.

Marie McGovern et al. (2002) Worm Breeder's Gazette "EGL-32 may be a Sperm Protein that Regulates Egg Laying though the TGF-b Pathway"

Cathy Savage-Dunn, Marie McGovern, Ling Yu

[Worm Breeder's Gazette, 2002]

A TGF-b related signaling pathway regulates dauer larval development and egg laying in C. elegans . Mutations in daf-7 ligand, daf-1 type I receptor, daf-4 type II receptor, and daf-8 and daf-14 Smads result in Dauer-constitutive/Egg-laying defective animals (1). These mutations are suppressed for both defects by mutations in daf-3 and daf-5 (2). We are interested in the role of this pathway in egg laying. Two other genes that affect egg laying, egl-4 and egl-32 , are also implicated in this pathway by their suppression by daf-3 and daf-5 (3). Mutations in egl-4 also have a weak Dauer-constitutive phenotype (4). Mutants of egl-32 are the only known mutants to be suppressed by daf-3 and daf-5 that are Egg-laying defective, but not Dauer-constitutive. Mutants of egl-32 retain about twice as many eggs as wild type animals: 30 instead of 14. Experimental evidence suggests that EGL-32 is a sperm protein that regulates egg laying in C. elegans . It has 3 close homologs, all of which are found in C. elegans . EGL-32 and its 3 closest homologs have been shown to be highly expressed in sperm (5). egl-32(n155) is a temperature sensitive mutation. Temperature shift assays reveal that L4 is the critical stage for EGL-32 inactivation. The L4 stage does not correspond to the time when eggs are laid, but when hermaphrodites produce sperm. Furthermore, the introduction of wild type sperm, by mating, into egl-32 animals results in a reduction in the number of eggs retained, and an increase in the number of eggs laid. The introduction of egl-32 sperm into wild type animals, by mating, causes wild type animals to lay fewer eggs and retain more eggs, many of which are at the comma stage or later. It has previously been described that meiotic maturation and ovulation in C. elegans is regulated partially by sperm (6). In female mutants, oocytes fail to undergo meiotic maturation and sheath cell contraction, which is necessary for ovulation, is irregular (7). Introduction of sperm, by mating, causes oocytes to complete maturation and sheath cells to begin to contract regularly, allowing ovulation. It has recently been found that the major sperm protein (MSP) supplies the signal for oocyte maturation and ovulation (6) in C. elegans . It is possible that a mechanism also exists to coordinate the time of fertilization with the time of egg laying to insure that eggs are not laid too soon or too late. It is possible that this mechanism functions either directly or indirectly through the TGF-b pathway. Further experiments will be done to test this hypothesis. 1.Savage-Dunn C, Cytokine and Growth Factor Reviews , 12, 2001 2.Patterson G.I.., Koweek A., Wong A., Liu Y. and Ruvkun G, Genes Dev. 11, 1997. 3.Trent C., Tsung N. and Horvitz H.R, Genetics. 104, 1983. 4.Daniels S.A., Ailion M., Thomas J.H. and Sengupta P, Genetics 156, 2000. 5.Reinke V., Smith H.E., Nance J., Wang J., Van Doren C., Begley R., Jones S.J.M., Davis E.B., Scherer S., Ward S., Kim S.K, Molecular Cell 6, 2000. 6.Miller M.A., Nguyen V.Q., Lee M., Kosinski M., Schedl T., Capriolo R.M. and Greenstein D. A, Science 291, 2001. 7.Strome S, J Cell Biol. 103, 1986.

Jones D et al. (1988) Worm Breeder's Gazette "non-coordinate expression of 16 kilodalton heat shock genes of C elegans."

Dixon DK, Jones D, Candido EPM

[Worm Breeder's Gazette, 1988]

Our studies of the genes encoding 16 kilodalton heat shock proteins ( hsp16's) in C. elegans have resulted in the characterization of two hsp16 loci. One locus, hsp16-1/48, encodes a pair of divergently transcribed genes, hsp16-1 and hsp16-48, which are exactly duplicated in an inverted orientation, forming a 4 kb inverted repeat containing four hsp16 genes. This locus has been assigned to a 368 kb contig which maps to chromosome V, by John Sulston and Alan Coulson. The other locus, which has not yet been mapped, codes for another pair of divergently arranged genes, hsp16-2 and hsp16-41. All four genes are under tight heat shock control. However, Northern blot analysis had shown that there is approximately 15-fold more mRNA from locus 16-2/41 than from 16-1/48, on a per gene basis (Jones et al., 1986). To explore the molecular basis for this difference in expression levels, we have used locus-specific hybridization probes to measure the mRNA content of embryos under a variety of heat shock conditions. Nuclear run-on assays demonstrate that the difference in expression of the hsp16 loci is not primarily due to transcription rates - the rate of hsp16-2 transcription in vitro is at most two-fold higher than that of hsp16-1. In contrast, the two loci differ dramatically in the duration of their active phase and in the kinetics of inactivation during a prolonged heat shock. When embryos are placed abruptly at 33 C, the induction of both hsp16-1 and hsp16-2 message is detectable within minutes, and both transcripts accumulate at similar rates. However, hsp16-1 transcript levels begin to decline in less than one hour, while those of hsp16-2 continue to increase rapidly for an additional 40 min. before beginning to decrease; under these conditions, the peak mRNA levels of the two genes differ by four fold. When embryos are heated gradually (1 C per 15 min.) to 33 C, the difference between loci is even more marked - the level of hsp16-2 message continues to increase at a high rate for more than 30 min. after hsp16-1 mRNA reaches a plateau, and the ratio of 16-2:16-1 message reaches 7:1, or 14:1 on a per gene basis. Although we cannot measure mRNA half lives directly due to the impermeability of the embryos to inhibitors of transcription, the levels of hsp16-1 and hsp16-2 message drop in parallel during recovery from heat shock, suggesting that the two messages have similar stabilities. Thus the major reason for the differences in expression of the two loci seems to be due to a difference in the shutdown phase of the heat shock response: although both loci are transiently activated by heat shock, locus 16-1/48 is more rapidly inactivated than locus 16-2/41. The hsp16 genes are inducible at all stages of development, but the induced levels of message are highest in L1 and lowest in adults. The difference in expression between the two loci is also greatest at L1. The thermotolerance of embryos was examined - abrupt 30 min. heat shocks had little effect on survival until 36 C was reached, and at 38 C the survival was only 30%. If embryos were raised gradually to 38 C, and kept at that temperature for 30 min., survival jumped to approximately 90%. Likewise, if embryos were pre-treated at 32 C for 30 min., then subjected to 38 C for 30 min., a greater than two-fold increase in survival was seen, relative to the abrupt heat shock. Pre- induction of the heat shock response thus greatly enhances survival in C. elegans as in other organisms, and the effect is most significant for a gradual temperature increase, a situation which is undoubtedly much more frequently seen by worms living outside in the real world.