Worm Breeder's Gazette 14(2): 76 (February 1, 1996)

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

mRNA expression patterns of a cell cycle regulator CDC25 in embryos

Mark A. Wilson, Renee V. Hoch, Andy Golden

ABL-Basic Research Program, FCRDC, Frederick, MD 21702

     Early in embryonic development, the 5 somatic founder cells, AB,
MS, E, C, and D, each have a distinct cell cycle periodicity.  The cell
cycle within a particular founder cell lineage is relatively synchronous
within that lineage, but asynchronous with the other founder cell
lineages.  The two germline cells, Z2 and Z3, are born early in
embryonic development, but do not divide during the remainder of
embryogenesis; they do proliferate extensively during larval and adult
development to generate over 1000 germline cells.  We have initiated a
project to examine the regulation of these cell cycles during C. elegans
development.  Our goals are to explore how developmental signals
regulate these various cell cycles as well as to determine whether
perturbation of the cell cycle can affect development.
     Biochemical and genetic experiments from other systems have
revealed that serine/threonine kinases regulate the progression from one
stage of the cell cycle to the next (i.e, CDC2, CDKs).  These kinases in
turn are regulated by phosphorylation and dephosphorylation.  In these
other systems, the regulated activity of CDC2 is required for G2 to
M-phase progression.  CDC2 is phosphorylated on a single tyrosine
residue by WEE1 to maintain it as an inactive kinase during much of the
cell cycle.  This phosphotyrosine residue and an adjacent
phosphothreonine residue are dephosphorylated by a dual-specificity
phosphatase CDC25 to activate CDC2 for mitosis.  We have begun to
examine the expression of these universally conserved cell cycle
regulators, namely, the tyrosine kinase, WEE1, and the dual-specificity
phosphatase, CDC25, during C. elegans development.  The cloning and
sequence of the C. elegans cdc2 gene, ncc-1, has been described
previously (Mori et al., Mol. Gen. Genet., 1994).  Seydoux and Fire
(Development, 1994) have shown that ncc-1 is a maternal message that
perdures ubiquitously throughout embryonic development.  
     A search of the available sequences from the Genome Consortium
revealed two candidate cdc25 genes (cosmids ZK637 and R05H5; Sulston et
al., Nature, 1992). The two cdc25 candidates have all of the
distinguishing features that identify them as CDC25 family
dual-specificity phosphatases.  Using SL1, SL2, and 3¹ UTR specific
primers and RT-PCR, we identified a SL1-trans-spliced cDNA for the cdc25
(ZK637) gene.  We used this cdc25 cDNA for RNA in situ studies to
examine the mRNA expression pattern in embryos.  The ZK637 cdc25 message
is maternal: it is abundant in the 1-cell embryo and is partitioned to
the anterior half of the embryo.  The message clearly partitions to AB
in the 2-cell embryo and then to the AB progeny in the next 2-3 AB
divisions.  The mRNA essentially is not detectable after the 16-24 cell
stage.  The message is absent or at very low levels in P1 and cells
derived from P1.  We have not observed any embryonic transcription of
this message by this in situ method.  We are currently cloning the R05H5
cdc25 cDNA for RNA expression analysis.
     We are also testing CDC25 antibodies to determine the protein
expression patterns of these cell cycle regulators throughout
embryogenesis and germline development.
     This research was sponsored by the National Cancer Institute, DHHS,
under contract with ABL.