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The group’s efforts are currently focused in two general
areas. The first deals with the posttranslational modification and the
subcellular localization of signal transduction proteins. The second
deals with stress signaling and the characterization of histidine
kinases.
Posttranslational modification and subcellular
localization of Ras proteins. It has been known for some time
that Ras undergoes posttranslational addition of lipids and this is
required for subcellular localization and the signaling functions of
Ras. We have been involved in this area for many years, beginning in
the mid-80’s with the demonstration that mutants in the CaaX box
affected prenylation and palmitoylation of yeast Ras. Furthermore, we
were the first to show that yeast and mammalian Ras proteins undergo
carboxymethylation. Despite extensive characterization of protein
prenylation and the development of farnesyltransferase inhibitors
(FTIs) by big pharma, many questions remain. Perhaps the biggest gap in
our knowledge was the failure to isolate the palmitoyl transferase for
Ras, or for that matter, any of the numerous palmitoylated cellular
proteins. In the last two years, our group succeeded in identifying the
yeast Ras palmitoyltransferase (Ras PAT), initially by a genetic
selection followed up by biochemical purification. The identification
of the Ras PAT provided the handle necessary to identify putative PATs
in yeast, flies, worms, mouse, and human. Some of the potential human
PAT genes have been linked to human diseases. For the foreseeable
future, we plan to capitalize on our identification of the first PAT.
We will continue to use yeast to address structure function issues and
to establish enzyme-substrate pairs for the yeast ORFs. We will
collaborate with labs involved with the characterization of mammalian
PATs.
Recent papers:
"Identification
of a Ras palmitoyl acyl transferase in Saccharomyces
cerevisiae." Lobo, S., Greentree, W.K., Linder, M.E., and
Deschenes, R.J. J.Biol. Chem. 277(43), 41268-41273
(2002).
"Erf4p
and Erf2p Form an Endoplasmic Reticulum-associated Complex
Involved in the Plasma Membrane Localization of Yeast Ras Proteins."
Zhao, L., Lobo, S., Dong, X., Ault, A., and Deschenes, R.J. J.Biol.
Chem. 277(51), 49352-9 (2002).
"New
insights into the mechanisms of protein palmitoylation."
Linder, M. and Deschenes, R.J. Biochemistry 42
(15):4311-20 (2003).
"Palmitoylation
and plasma membrane localization of Ras2p by a nonclassical trafficking
pathway in Saccharomyces cerevisiae."
Dong, X.,Mitchell, D.A., Lobo, S., Zhao, L., Bartels, D.J.,and
Deschenes, R.J. Mol Cell Biol. Sep;23(18):6574-84. (2003)
"Model
organisms lead the way to protein palmitoyl-transferases."
Linder, M.E. and Deschenes, R.J. J. Cell Sci. 117:521-526
(2004).
Genetic dissection of two-component regulators in yeast Recent
characterization of the osmotic response pathway in S. cerevisiae has
uncovered a complex pathway consisting of two signal transduction
modules. Changes in osmotic balance are sensed by a sensor-kinase,
Sln1p, which undergoes autophosphorylates a conserved histidine. That
phosphoryl group is then transferred to a conserved aspartate in the
Sln1 receiver domain, and then to a histidine in the phosphorelay
molecule, Ypd1p and finally to an aspartate on the independent receiver
domain of the response regulator, Ssk1p. The HOG1 MAP kinase pathway is
kept in the inactive state under normal osmotic conditions by virtue of
Sln1p and thus Ssk1p phosphorylation, since activation of the HOG1 MEK
kinase requires a dephosphorylated form of Ssk1p. We are interested in
this pathway for several reasons. First, little is known about how
osmotic stress is sensed in any system and yeast affords a nice
opportunity to uncover basic principles. Second, histidine kinases are
an interesting subset of protein kinases that has been somewhat
overlooked in favor of Tyr and Ser/Thr kinases. Finally, two component
histidine protein kinases are potential antifungal drug targets.
Currently, systemic fungal infections are difficult to treat because of
drug toxicity and the increasing emergence of resistance to the small
group of available antifungals. Therefore, a better understanding of
the function of fungal histidine kinases and associated two component
like proteins could lead to improved antifungal drugs with reduced
lower toxicity. This project is a collaborative effort between my lab
and Dr. Jan Fassler at the University of Iowa.
Recent papers:
"The
cytoplasmic linker region of the Sln1 histidine kinase forms
a coiled-coil important in regulation of kinase activity."
Tao, W., Malone, C.L., Ault, A.D., Deschenes, R.J., Fassler, J.S. Mol.
Micro. 43(2):459-73 (2002.
"The
eukaryotic two-component histidine kinase
Sln1p regulates OCH1 via the transcription factor, Skn7p."
Li, S., Dean, S., Li, Z., Horecka, J., Deschenes, R.J., and Fassler,
J.S., (2002) Mol Biol Cell
13(2):412-24 (2002).
"Ypd1p,
a Saccharomyces cerevisiae Histidine Phosphotransferase, shuttles
between
the nucleus and cytoplasm for SLN1-dependent phosphorylation of Ssk1p
and Skn7p."
Lu, M.-Y., Deschenes, R.J., and Fassler, J.S. Euc. Cell. 2:
1304-14 (2003).
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