Table 1. Yeast strains used in this study Strain
Genotype
Source
S18-1D
MATα his3 leu2 ura3 trpl ade8 tpk1w tpk2::HIS3
Ref.3
tpk3::TRP1 TP408a
S18-1D krh1::URA3
This study
TP626a,b
S18-1D krh1::URA3 krh2::KanMX4
This study
RS13-58A-1
MATa his3 leu2 ura3 trpl ade8 tpk1w tpk2::HIS3
Ref.3
tpk3::TRP1 bcy1::LEU2 TP415a
RS13-58A-1 krh1::URA3
This study
TP627a,b
RS13-58A-1 krh1::URA3 krh2::KanMX4
This study
BY4742
MATα leu2 lys2 ura3 his3
Ref.4
Y10351
MATα leu2 lys2 ura3 krh1::KanMX4
Ref.5
Y11668
MATα leu2 lys2 ura3 his3 krh2::KanMX4
Ref.5
Y00351
MATa leu2 met15 ura3 his3 krh1::KanMX4
Ref.5
TP261d
MATα krh1::KanMX4 krh2::KanMX4
This study
Y10152
MATα leu2 lys2 ura3 his3 gpa2::KanMX4
Ref.5
TP675c
TP261 gpa2::LEU2
This study
TP676k
TP261 GPA2(Q300L)::LEU2
This study
TP677k
BY4742 GPA2(Q300L)::LEU2
This study
TP380a,e,f
MATa/α cyr1::KanMX4/CYR1 pde2::URA3/PDE2
This study
krh1::LEU2/KRH1 krh2::HIS3/KRH2 TP390g
cyr1::KanMX4 pde2::URA3 krh1::LEU2 krh2::HIS3
This study
TP420e,f
MATa/α cyr1::KanMX4/CYR1 pde2::URA3/PDE2
This study
TP433h
MATα cyr1::KanMX4 pde2::URA3
This study
TP522a,e,f,i
MATa/α cyr1::KanMX4/CYR1 pde2::URA3/
This study
pde2::URA3 krh1::HIS3/KRH1 RAS2(G19V)::LEU2 TP519j
cyr1::KanMX4 pde2::URA3 RAS2(G19V)::LEU2
This study
TP467e,f,k
MATa/α cyr1::KanMX4/CYR1 pde2::URA3/PDE2
This study
GPA2(Q300L)::LEU2 TP506l
cyr1::KanMX4 pde2::URA3 GPA2(Q300L)::LEU2
This study
TP538a,m
MATa/α tpk1::KanMX4/tpk1::KanMX4
This study
tpk2::KanMX4/tpk2::KanMX4 tpk3::LEU2/TPK3 yak1::URA3/YAK1 TP543n
MATα tpk1::KanMX4 tpk2::KanMX4 tpk3::LEU2
This study
yak1::URA3 TP544n
MATα tpk1::KanMX4 tpk2::KanMX4 tpk3::LEU2
This study
yak1::URA3 TP561a,m
MATa /alpha tpk1::KanMX4/tpk1::KanMX4
This study
tpk2::KanMX4/tpk2::KanMX4 tpk3::LEU2/TPK3 yak1::URA3/YAK1 krh1::HIS3/KRH1 krh2::HIS3/KRH2 TP571o
MATα tpk1::KanMX4 tpk2::KanMX4 tpk3::LEU2
This study
yak1::URA3 krh1::HIS3 krh2::HIS3 TP572o
MATa tpk1::KanMX4 tpk2::KanMX4 tpk3::LEU2
This study
yak1::URA3 krh1::HIS3 krh2::HIS TP573a,p
MATa yak1::URA3 HIS3 LEU2
This study
TP575a
MATα krh1::LEU2 krh2::HIS3 yak1::URA3
This study
TP488p,q
MATa HIS3 LEU2
This study
TP446a
MATa krh1::LEU2 krh2::HIS3
This study
TP448a,m
MATa/α tpk2::KanMX4/TPK2 tpk3::LEU2/TPK3
This study
krh1::LEU2/KRH1 krh2::HIS3/KRH2 TP462r
MATa tpk2::KanMX4 tpk3::LEU2 krh1::LEU2
This study
krh2::HIS3 TP449a,m
MATa/α tpk1::KanMX4/TPK1 tpk3::LEU2/TPK3
This study
krh1::LEU2/KRH1 krh2::HIS3/KRH2 TP461s
MATα tpk1::KanMX4 tpk3::LEU2 krh1::LEU2
This study
krh2::HIS3 TP450a,m
MATa /α tpk1::KanMX4/TPK1 tpk2::KanMX4/TPK2
This study
krh1::LEU2/KRH1 krh2::HIS3/KRH2 TP465t
MATa tpk1::KanMX4 tpk2::KanMX4 krh1::LEU2
This study
krh2::HIS3 TP482a,m,p
MATa tpk2::KanMX4 tpk3::LEU2 HIS3
This study
TP491a,m,p
MATα tpk1::KanMX4 tpk3::LEU2 HIS3
This study
TP486m,p,q
MATa tpk1::KanMX4 tpk2::KanMX4 LEU2 HIS3
This study
MV600u
MATa KRH1(HA)3::URA3
This study
MV601v
MATa HIS3MX6::P(GAL1)(HA)3CYR1
This study
MV602v
MATa HIS3MX6::P(GAL1)GST-GPA2
This study
Y00152
MATa leu2 met15 ura3 his3 gpa2Δ::KanMX4
Ref.5
MV603k
MATa gpa2Δ::KanMX4 GPA2(Q300L)::LEU2
This study
MV604k,v
MATa gpa2Δ::KanMX4 HIS3MX6::P(GAL1)GST-
This study
GPA2(Q300L)::LEU2
MV605u,v
MATa HIS3MX6::P(GAL1)GST-GPA2
This study
KRH1(HA)3::URA3 MV606k,u,v
MATa gpa2Δ::KanMX4 HIS3MX6::P(GAL1)GST-
This study
GPA2(Q300L):: LEU2 KRH1(HA)3::URA3 MV607w
MATa [YEpGFP-HA-BCY1]
This study
MV608u,w
MATa [YEpGFP-HA-BCY1] KRH1-HA3::URA3
This study
pJ69-4A
MATa leu2-3,112 ura3-52 trp1-901 his3-200 gal4Δ
Ref.6
gal80Δ GAL-ADE2 lys2::GAL1-HIS3 met2::GAL7LacZ TP636a
pJ69-4A krh1::URA3
This study
TP637a,b
pJ69-4A krh1::URA3 krh2::KanMX4
This study
TP608a,m,x
MATa/α tpk1::KanMX4/tpk1::KanMX4
This study
tpk2::KanMX4/tpk2::KanMX4 tpk3::LEU2/TPK3 krh1::HIS3/KRH1 krh2::HIS3/KRH2 mouseCα2::URA3 TP665y
MATa tpk1::KanMX4 tpk2::KanMX4 tpk3::LEU2
This study
mouseCα2::URA3 TP667y
MATa tpk1::KanMX4 tpk2::KanMX4 tpk3::LEU2
This study
krh1::HIS3 krh2::HIS3 mouseCα2::URA3 SP1
MATa leu2 his3 trp1 ura3 ade8 can1
Ref.7
MWY63
MATα leu2 his3 trp1 ura3 ade8 tpk1::URA3 tpk2-
Ref.8
63(ts) tpk3::TRP1 bcy1::LEU2 MV620a,b
MWY63 krh1::KanMX4 krh2::HIS3
This study
TP701i
S18-1D RAS2(G19V)::LEU2
This study
a
Plasmids of the pRS series were used to generate PCR disruption cassettes (4).
b
Genomic DNA of Y00351 or Y11668 was used as a template for PCR of a
krh1::KanMX or krh2::KanMX4 disruption cassette, respectively. c
A linear fragment of pUC19-gpa2::LEU2 was used as disruption cassette.
d
Tetrad segregant of a diploid strain generated by crossing Y00351 with Y11668.
e
A linear fragment of ppde2::URA3 was used as disruption cassette (9).
f
Disruptions were made in strain Y26773 (5).
g
Tetrad segregant of strain TP380.
h
Tetrad segregant of strain TP420.
i
YIplac128-RASG19V was cut with PstI for direct integration at the RAS2 promoter
locus. j
Tetrad segregant of strain TP522.
k
YIplac128-GPA2Q300L was cut with MluI for direct integration at the GPA2 promoter
locus. l
Tetrad segregant of strain TP467.
m
tpk1::KanMX4 and/or tpk2::KanMX4 derived from YO1261 and/or Y11089
respectively (5). n
Tetrad segregant of strain TP538.
o
Tetrad segregant of strain TP561.
p
pJJ215 was cut with NsiI for direct integration at the HIS3 terminator locus.
q
pYIplac128 was cut with PflMI for direct integration at the LEU2 promoter locus.
r
Tetrad segregant of strain TP448.
s
Tetrad segregant of strain TP449.
t
Tetrad segregant of strain TP450.
u
pYX012/KRH1 was digested with SphI to direct integration of P(TPI1)-KRH1-HA3
at the TPI1 promoter locus. v
The HIS3MX6GAL1-GST or HIS3MX6GAL1-HA3 cassette (10) was used to replace
the endogenous promoter with GAL1 and an in-frame N-terminal GST fusion. w
Constructed by direct transformation with plasmid YEplac181-GFP-HA-BCY1
(provided by G. Griffioen, KU Leuven). x
YIp-PromTPK1-HA3-mouseCα2-TermTPK1 was cut with AgeI for direct integration at
the TPK1 promoter locus. y
Tetrad segregant of strain TP608.