Biology Reference
In-Depth Information
limit the cAMP signal is, in part, a function of PDE activity (Ma et al.
1999
; Nikawa
et al.
1987a
; Wang et al.
2005
).
Most of the components of the
S. pombe
cAMP pathway have been identified
from mutant strains that are defective in glucose repression of transcription of the
fbp1
gene, which encodes the gluconeogenic enzyme fructose-1,6-bisphosphatase
(Hoffman and Winston
1990
,
1991
). Key to these genetic selections is a reporter
construct in which the
fbp1
promoter drives expression of the
ura4
OMP decarbox-
ylase gene. The
ura4
gene is a selectable marker as its expression is required for
uracil biosynthesis, which is needed for growth in medium lacking uracil. It is also a
counterselectable marker, as the pyrimidine analog 5-fluoroorotic acid (5FOA) is
toxic to cells that express it. Normally,
S. pombe
represses
fbp1-ura4
transcription
in glucose-rich medium, preventing growth in medium lacking uracil while allow-
ing growth in 5FOA medium. Mutants defective in glucose/cAMP signaling, which
is required to activate the cAMP-dependent protein kinase (PKA), constitutively
transcribe the
fbp1-ura4
construct so as to confer growth on medium lacking uracil,
as well as 5FOA-sensitivity (5FOA
S
). These strains carry mutations that affect the
Git3 G protein-coupled receptor (Welton and Hoffman
2000
), any subunit of the
heterotrimeric G protein composed of Gpa2 (Nocero et al.
1994
), Git5 (Landry et al.
2000
), and Git11 (Landry and Hoffman
2001
), the Git2/Cyr1 adenylyl cyclase
(Hoffman and Winston
1991
), the Git1 adenylyl cyclase-binding protein (Kao
et al.
2006
), Hsp90 (Alaamery and Hoffman
2008
), the Git7 Hsp90 co-chaperone
(Schadick et al.
2002
), or the Pka1 PKA catalytic subunit (Jin et al.
1995
) (Fig.
1
).
Fig. 1 The
S. pombe
glucose/cAMP signaling pathway presented using the ProteinLounge
Pathway Builder Tool
