Genetic Approaches
Identification of genes with cell autonomous functions in C. elegans distal tip cell migration and gonadogenesis.
1Dept. of Molecular Biology, Princeton University, Princeton, NJ 08544 2Dept. of Biochemistry, University of Wisconsin, Madison, WI 53706
A C. elegans genome-wide screen identified 99 genes as required for distal tip cell (DTC) migration during gonadogenesis (Cram et al., 2006). To determine which of the 99 genes affect DTC migration cell-autonomously, we used a C. elegans strain that allows RNAi to have an effect mainly in the two DTCs. 28 of the 99 genes caused DTC migratory defects in a primary screen. We conclude that these genes act cell autonomously. The types of gonad defects were scored in a secondary screen and the percentages in each of the three types are shown in the table. Further information about the screen follows the table.
Reference: Cram, E. J., H. Shang, and J. E. Schwarzbauer (2006) A systematic RNA interference screen reveals a cell migration gene network in C. elegans. J. Cell Sci. 119, 4811-4818.
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| Distal Tip Cell Specific C. elegans Genes Affecting Migration | |||||||
| Gene | Description/Function | Parental P0 defect1 | Distribution (percent)2 | Progeny F1 defect1 | Distribution2 (percent) | Decreased F1 progeny3 | |
| 1 | Gene: (B0511.8) Gene Pair ID CMKB Entrez Gene | Temporarily Assigned Gene name AhringerAutoClass: Signaling Function: Metabolism InterPro: Stuctural constituent of ribosome | (N=19) 0 | (N=84) 0 | |||
| 2 | Gene: (C01G8.5) Gene Pair ID CMKB Entrez Gene | Ezrin/Radixin/Moesin AhringerAutoClass: Unknown Function: Cell Architecture InterPro: Cytoskeletal protein binding | (N=12) 0 | (N=94) 0 | |||
| 3 | Gene: (C02C6.1) Gene Pair ID CMKB Entrez Gene | DYNamin related AhringerAutoClass: Cell Architecture Function: Cell Architecture InterPro: GTPase | (N=20) 0.15 | Type 1: 33 Type 2: 30 Type 3: 37 | (N=20) 0.7 | Type 1: 42 Type 2: 17 Type 3: 42 | |
| 4 | Gene: (C02F4.1) Gene Pair ID CMKB Entrez Gene | CEll Death abnormality AhringerAutoClass: Cell Architecture Function: Cell Architecture InterPro: *Small GTPase regulatory/interacting protein | (N=72) 0.01 | (N=79) 0.03 | |||
| 5 | Gene: (C06G3.10) Gene Pair ID CMKB Entrez Gene | Conserved Oligomeric Golgi (COG) Component AhringerAutoClass: Unknown Function: Unknown InterPro: | (N=42) 0 | (N=59) 0 | |||
| 6 | Gene: (C09G12.8) Gene Pair ID CMKB Entrez Gene | CEll Death abnormality AhringerAutoClass: Signaling Function: Signaling InterPro: GTPase | (N=110) 0 | (N=142) 0.01 | |||
| 7 | Gene: (C09H10.7) Gene Pair ID CMKB Entrez Gene | C09H10.7 AhringerAutoClass: Unknown Function: Unknown InterPro: | (N=20) 0.35 | Type 1: 32 Type 2: 14 Type 3: 54 | (N=20) 0.15 | Type 1: 30 Type 2: 11 Type 3: 60 | |
| 8 | Gene: (C15F1.3) Gene Pair ID CMKB Entrez Gene | TRAnsformer : XX animals transformed into males AhringerAutoClass: Unknown Function: Unknown InterPro: *Transmembrane receptor | ND | ||||
| 9 | Gene: (C16C10.6) Gene Pair ID CMKB Entrez Gene | CCDC (human Coiled Coil Domain Containing) homolog AhringerAutoClass: Unknown Function: Unknown InterPro: | (N=13) 0 | (N=26) 0 | |||
| 10 | Gene: (C24D10.4) Gene Pair ID CMKB Entrez Gene | C24D10.4 AhringerAutoClass: Unknown Function: Unknown InterPro: | (N=77) 0.03 | (N=57) 0.09 | |||
| 11 | Gene: (C27H6.2) Gene Pair ID CMKB Entrez Gene | RUVB (recombination protein) homolog AhringerAutoClass: Unknown Function: DNA Binding InterPro: ATP binding DNA helicase | (N=57) 0.04 | (N=11) 0.18 | |||
| 12 | Gene: (C28C12.8) Gene Pair ID CMKB Entrez Gene | Helix Loop Helix AhringerAutoClass: Transcription Factor Function: Transcription Factor InterPro: | (N=58) 0.01 | Type 1: 38 Type 2: 19 Type 3: 44 | (N=19) 0.5 | Type 1: 29 Type 2: 12 Type 3: 59 | |
| 13 | Gene: (C36E8.5) Gene Pair ID CMKB Entrez Gene | Tubulin, Beta AhringerAutoClass: NULL Function: Cell Architecture InterPro: GTPase/Structural molecule | (N=55) 0.67 | Type 1: 5 Type 2: 35 Type 3: 60 | (N=20) 0.85 | Type 1: 17 Type 2: 28 Type 3: 56 | |
| 14 | Gene: (C37C3.6) Gene Pair ID CMKB Entrez Gene | PaPiliN (Drosophila ECM protein) homolog AhringerAutoClass: Unknown Function: ECM InterPro: Metalloendopeptidase/serine protease inhibitor | (N=37) 1 | Type 1: 71 Type 2: 11 Type 3: 17 | (N=0) 0 | ||
| 15 | Gene: (C43H6.9) Gene Pair ID CMKB Entrez Gene | GLutamate Receptor family (AMPA) AhringerAutoClass: Neuro Function: Other InterPro: Glutamate gated ion channel | (N=80) 0.03 | (N=76) 0.03 | |||
| 16 | Gene: (C47B2.3) Gene Pair ID CMKB Entrez Gene | C47B2.3 AhringerAutoClass: NULL Function: Cell Architecture InterPro: GTPase/Structural molecule | (N=15) 0.6 | Type 1: 7 Type 2: 67 Type 3: 27 | (N=20) 0.8 | ||
| 17 | Gene: (C47E12.4) Gene Pair ID CMKB Entrez Gene | inorganic PYroPhosphatase AhringerAutoClass: Metabolism Function: Metabolism InterPro: Magnesium ion binding pyrophosphatase | (N=94) 0 | (N=41) 0.1 | |||
| 18 | Gene: (C47E8.7) Gene Pair ID CMKB Entrez Gene | UNCoordinated AhringerAutoClass: Cell Architecture Function: Cell Architecture InterPro: *Cell adhesion constituent | (N=65) 0.05 | (N=7) 0 | |||
| 19 | Gene: (C53B4.1) Gene Pair ID CMKB Entrez Gene | C53B4.1 AhringerAutoClass: Small Molecule Transport Function: Other InterPro: Transporter | (N=48) 0.02 | (N=51) 0.16 | |||
| 20 | Gene: (C56C10.8) Gene Pair ID CMKB Entrez Gene | Inhibitor of Cell Death AhringerAutoClass: Synthesis Function: Transcription Factor InterPro: | (N=7) 0 | (N=26) 0 | |||
In the original screen that identified 99 DTC migration genes, rrf-3(pk1426) worms were subjected to RNA interference (RNAi) via bacteria expressing doubled-stranded RNA (Cram et al., 2006). A caveat to this approach is that all tissues have the potential to be affected by RNAi; DTC migration might be affected by RNAi knockdown in tissues other than the gonad. To determine which of the 99 genes affect DTC migration cell autonomously, we took advantage of using lag-2p::GFP; rde-1(ne219); lag-2p::rde-1 strain of C. elegans (generated by Dr. Dana Byrd and Dr. Judith Kimble) that allows RNAi to have an effect mainly in the two DTCs. We applied the same 2-step microscopy approach as Cram et al. and examined DTC migratory defects resulting from RNAi knockdown of each of the 99 genes. We found that 28 of these genes caused DTC migratory defects in ≥ 30% of treated nematodes in the primary screen. We conclude that these genes act cell autonomously. The types of gonad defects were scored in the secondary screen, as defined by Cram et al.., and the percentages in each of the three types are shown in the table. Also noted in the table is whether the number of F1 progeny was reduced, which may be caused by morphological defects of the gonad that impact embryogenesis. RNAi of the remaining 71 genes had no significant effect on DTC migration. However, we cannot conclude that these genes are not acting cell autonomously because the absence of a defect could result from incomplete knockdown associated with feeding C. elegans RNAi bacteria.
Notes:
ND -- data not determined due to no RNAi bacterial growth or gene lethality.
1 -- Primary screen: assayed for presence of clear patch phenotype under light microscopy.
2 -- Secondary screen: categorized DTC migration defects under DIC microscopy.
3 -- Observed no/reduced F1 progeny in either screen.
Type 1 -- Ventralized gonad arms.
Type 2 -- Gonad arms with supernumary turns and inappropriate dorsal turns.
Type 3 -- Dorsal gonadal defects.