Program of the 2001 International Worm Meeting - Sternberg Lab ...

181
181. Two enhancers of raf, eor-1 and
eor-2, function in parallel to sur-2,
lin-25, and lin-1 to positively regulate
Ras signaling
Robyn M. Howard, Meera V.
Sundaram
University of Pennsylvania School of Medicine,
Philadelphia, PA 19104
In C. elegans, the Ras/Raf/MEK/ERK signaling
pathway is required for many processes. To
identify Ras pathway modifiers, our lab
conducted a screen for mutations that alone
cause Ras-like defects at low penetrance but that
in combination with a weak lin-45raf allele,
ku112, cause Ras-like defects at high penetrance
(see abstract by Rocheleau et. al.). This screen
has isolated loss-of-function mutations in two
previously uncharacterized enhancers of raf,
eor-1 and eor-2, each defined by several alleles.
eor-1 and eor-2 do not show genetic interactions
with each other but show strong genetic
interactions with sur-2, lin-25, and lin-1, genes
encoding transcriptional regulators that
genetically act downstream of mpk-1. Our data
suggest that eor-1 and eor-2 function together to
positively regulate Ras signaling at a similar
step but in parallel to sur-2, lin-25, and lin-1.
eor-1/eor-2 and sur-2/lin-25 are redundantly
required for multiple outputs of Ras signaling.
We propose that these two sets of genes may be
the key positively acting downstream
transcriptional regulators that are controlled by
Ras signaling in several tissues. Consistent with
eor-1 and eor-2 acting as transcriptional
regulators, eor-1 encodes a BTB/Zinc finger
protein, similar to human promyelocytic
leukemia zinc finger protein (hPLZF). PLZF
can bind corepressor complex components and
mouse knockouts of PLZF show decreased Hox
gene expression. We are testing whether EOR-1
binds corepressor complex components and
whether eor-1 and eor-2 regulate Hox gene
expression.
182. Protein Phosphatase 2A is a
positive regulator of the Ras
pathway.
Gautam Kao, Meera Sundaram
Dept. of Genetics, Univ. of Pennsylvania
Medical School, Philadelphia PA 19104, USA
182
sur-6 is a positive regulator of the ras pathway
and acts between ras and raf. It encodes the
PR55 regulatory subunit of the heterotrimeric
protein phosphatase 2A (PP2A). By several
genetic interaction criteria, sur-6 mutants are
very similar in their behavior to those of another
positive regulator ksr-1 that encodes a raf-like
kinase. We present data to show that the protein
phosphatase 2A (PP2A) holoenzyme is a
positive modulator of the Ras pathway.
Depending on the cellular context, the PR55
regulatory subunit can potentiate or inhibit the
activity of the PP2A holoenzyme. We wished to
understand whether SUR-6 activated or
inhibited the catalytic activity of PP2A in the
context of vulval fate specification. Work by the
Kohara and Baillie labs showed that let-92
encodes the catalytic subunit of PP2A and that
the strong let-92(s504) mutation causes worms
to die as young larvae before vulval
development commences. We have shown that
let-92(s504) is a strong dominant
(haplo-insufficient) enhancer of two let-60(dn)
alleles, n2031 and sy100, for their vulval
induction defects. The strong lin-45 Raf(oz166
lf) mutation has a similar dominant genetic
interaction with let-60(n2031). Additionally,
let-92 soaking RNAi causes a weak vulvaless
phenotype in a sensitized genetic background.
Therefore reducing let-92 function impairs the
ability of VPCs to adopt vulval fates. Finally
let-92(s504) is also a dominant lethal in a sur-6
background, indicating that sur-6 mutants have
reduced PP2A activity. Taken together these
data imply that the catalytic activity of PP2A is
positive regulator of the Ras-MAPK
phosphorylation cascade and that its action is
potentiated by SUR-6. Its likely role would be
to remove inhibitory phosphates that repress the
activity of other positive regulators of the
pathway.