Figure 1.
mpk-1 Expression in the C. elegans Germline (A) Schematics of
mpk-1a and
mpk-1b mRNAs. Box, exon; connecting line, intron; ATG, initiation codon; TAG, termination codon. Below schematics: thick bars, extent of probes used for in situ hybridization; arrows, primer pairs used for RT-PCR. (B) Semiquantitative RT-PCR of RNA prepared from adult hermaphrodites that either had an essentially normal germline [
glp-1(
q224) grown at 15 C], or had virtually no germline [
glp-1(
q224) grown at 25 C].
unc-54 was used as a control. (C) Western blot. MPK-1A protein is 45 kDa, MPK-1B is 55 kDa, and alfa-TUB is alfa-tubulin. Proteins were extracted from adult hermaphrodites that were either wild-type (wt),
glp-1(
q224) grown at 25 C (GL-), or
mpk-1(
ga117) putative null homozygotes [
mpk-1(0)]. (D-F) In situ analysis of dissected adult hermaphrodite germlines. (D) Total
mpk-1 RNA was assessed using the
mpk-1ab antisense probe shown in (A). (E)
mpk-1b RNA was assessed using an isoform-specific antisense probe shown in (A). (F) Negative control, using an
mpk-1b-specific sense probe. (G-L) Immunocytochemistry of dissected adult hermaphrodite germlines. All were stained using both MPK-1 antibodies (G, I, K) and DAPI (H, J, L). Distal end, arrowhead; dotted lines, boundaries between regions of germline maturation [MR (mitotic region), TZ (transition zone), PR (pachytene region), OO (oocytes), SP (sperm)]; PEX (pachytene exit defect). (G, H) Same wild-type germline. (I, J) Same
mpk-1(0) germline. (K, L) Same
mpk-1b(RNAi) germline.