Reagents

Scientific image of

Parkhurst Lab reagents available from the Development Studies Hybridoma Bank

AntigenMonoclonal LineSpeciesReference 
Pavarotti (Pav)O8fly20RRID: AB_2936338
Pavarotti (Pav)L24fly20RRID: AB_2936339   
Rho1P1D9fly3RRID: AB_528263 
WashP3H3fly8, 12RRID: AB_2618389
WashP4C9fly12RRID: AB_2618391 
WASPP5E1fly12RRID: AB_2618392 
WASPP3B1fly12RRID: AB_2618388 
SCARP1C1fly12RRID: AB_2618386 
SCARP1C8fly12 
WhamyP4A8fly12RRID: AB_2618390 
WhamyP1D1fly12 
p120cateninP1B2fly3, unpublishedRRID: AB_10660833 
p120cateninP4B2fly3, unpublishedRRID: AB_2088073 
dSir2P2E2fly4, unpublishedRRID: AB_1553776 
dSir2P4A10fly4, unpublishedRRID: AB_1553778 

Parkhurst Lab reagents available from the Bloomington Drosophila Stock Center

 

Stock #

Genotype

Allele type | insertion chromosome

Reference

wimp (RpII140 – RNA Polymerase II subunit)

5874

wimp, rucuca/TM3, Sb

antimorphic allele

1

Rho1 GTPase

9477

w*; b pr cn Rho1[1B] px sp/CyO

null allele

2,3,5

Cdc42  [R186C point mutation]

[Cdc42 point mutation (Cdc42(R186C)) under control of the UASp promoter (inducible)]
[This is a dominant negative Cdc42 mutation that gives similar phenotypes in flies to that observed in the equivalent human mutations.]

90925

w*  P{w+, UASp-Cdc42(R186C)}11

X

15

90926

w*; P{w+, UASp-Cdc42(R186C)}26/CyO

2nd

15

90927

w*; P{w+, UASp-Cdc42(R186C)}37

3rd

15

washout  (Wiskott-Aldrich Syndrome family protein)

79220

w*; wash[Delta185]

null allele,background lethal removed

14

28285

w*; wash[Delta185]/CyO

null allele

6,9


Sqh-mScarlet

[This is a CRISPR knock-in of mScarlet-i into the Spaghetti squash locus (C-terminal fusion)]

94929

TI{TI}sqhmScarlet-I ; MKRS/TM6B

CRISPR knock-in (X chr)

19

sqh gRNA

[gRNA for sqh knock-in into pCFD5 vector]

94930

w*;  P{U6:3-sqh.gRNA.pCFD5}attp2

gRNA insertion on 3rd

19

degringolade

35525

w*; dgrn[DK]/TM3, Sb

null allele

9

"Red" balancers (sChFP : sqh driven, ChFP protein)

35522

FM7a, P{w+; sChFP}

X

11

35523

w*; Sco/CyO, P{w+, sChFP}

2nd

11

35524

w*; Gl/TM3, Sb P{w+, sChFP}    

3rd

11

photoconvertible actin binding domain

[sM3MCA : sqh driven, Maple3 fluorescent protein, moesin α-helical-coiled and actin binding site]  

94924

w*;  P{w+, sM3MCA}12 

X

19

94925

w*;  P{w+, sM3MCA}20/CyO; MKRS/TM3,Ser

2nd

19

94926

w*;  Kr/CyO; P{w+, sSM3MCA}33 

3rd

19

actin binding domain

[sStMCA : sqh driven, Scarlet fluorescent protein, moesin α-helical-coiled and actin binding site]
[This Scarlet version gives much brighter fluorescence and less background than mCherry or Kate2. This reporter also recognizes nuclear actin.]

90928

w*  P{w+, sStMCA}10

X

16

90929

w*;  P{w+, sStMCA}20

2nd

16

90930

w*;  P{w+, sStMCA}20

3rd

16

actin binding domain

(sChMCA : sqh driven, ChFP protein, moesin α-helical-coiled and actin binding site)

35519

 w* P{w+, sChMCA}#12

X

10

35520

w*; P{w+, sChMCA}#22

2nd

10

35521

w*; P{w+, sChMCA}#31

3rd

10

actin binding domain

(sK2MCA: sqh driven, Kate2 fluorescent protein, moesin α-helical-coiled and actin binding site)
(Kate2 is supposed to be far red.  It isn’t in flies, but it is better separated from GFP than CherryFP (giving less background fluorescence)).

76268

w* P{w+, SK2MCA}#12

X

14

76269

w*;  P{w+, sK2MCA}#12

2nd

14

76270

w*;  P{w+, sK2MCA}#12

3rd

14

actin (inducible RFP fusion protein)

24777

P{UASp-RFP.actin}13, w*

X

7

24778

w*; P{UASp-RFP.actin}29

2nd

7

24779

w*; P{UASp-RFP.actin}38

3rd

7

GCaMP6s

[This is a constitutive version of the GCaMP6s calcium reporter that works very well in embryos.]

91362

w[*]; P{w[+mC]=sqh-GCaMP6s]101.1 sqh-GCaMP6s]101.2

X    (two copies of transgene)

17

91363

w[*]; P{w[+mC]=sqh-GCaMP6s}20

2nd  (one copy of transgene)

17

91364

w[*]; P{w[+mC]=sqh-GCaMP6s]305.1 sqh-GCaMP6s]305.2

3rd  (two copies of transgene)

17

GCaMP6s

[This is an inducible (UAS) version of the GCaMP6s calcium reporter in the pUASp vector (works well in the germline and embryo).]

91365

w[*]; P{w[+mC]=UASp-GCaMP6s}20/CyO

2nd

 

91366

w[*]; P{w[+mC]=UASp-GCaMP6s}30/TM3, Sb

3rd

 

Sep1 RNAi

(Express shRNA for Sep1 under UASz control)

604937

w* ; P{y[+t7.7]=UASz-Sep1-shRNA }JK22C

2nd

21

604938

w*;;  M{UASz-Sep1-shRNA }ZH-86Fb

3rd

21

AnxB10-mScarlet

(This is a CRIMIC knock-in of mScarlet-i (StFP) into the AnxB10 locus)

605570

y[1] w[*] TI{CRIMIC.StFP}AnxB10[CR00582-StFP]

X

22

sfGFP-AnxB10

(sfGFP-AnxB10 fusion protein under control of sqh promoter)

605571

w*, P{w[+mC]=sqh-sfGFP-AnxB10}10

X

22

605572

w*;  P{w[+mC]=sqh-sfGFP-AnxB10}20; MKRS/TM6B

2nd

22

605573

w*;;  P{w[+mC]=sqh-sfGFP-AnxB10}30

3rd

22

StFP-AnxB11

(mScarlet-i (StFP)-AnxB11 fusion protein under control of endogenous AnxB11 promoter)

605574

w*, P{StFP-AnxB11 }10

X

22

605575

w*; P{StFP-AnxB11 }20

2nd

22

605576

w*;; P{StFP-AnxB11 }31

3rd

22

 

AnxB11 RNAi

(Express shRNA for AnxB11 under Walium22 control)

605568

w*;;  M{UAS-AnxB11-shRNA1 }ZH-86Fb

3rd

22

 

605569

w*;;  M{UAS-AnxB11-shRNA2 }ZH-86Fb

3rd

22

 

StFP-8PM

(Plasma membrane specific reporter.  mScarlet-i (StFP) fused to prenylated octavalent that associates to plasma membrane under control of Ubiquitin promoter)

605577

w*, P{w[+mC]=Ubi-StFP-8PM }10

X

22

605578

w*;  P{w[+mC]= Ubi-StFP-8PM }25

2nd

22

605579

w*;;  P{w[+mC]= Ubi-StFP-8PM }31

3rd

22

StFP-Lact-C2

(PS lipid biosensor.  mScarlet-i (StFP) fused to Lact-C2 domain that associates to phosphatidylserine under UASz control)

605580

w*;;  M{UASz-StFP-Lact-C2 }ZH-86Fb

3rd

22

PASS-sfGFP

(PA lipid biosensor.  sfGFP fused to PASS domain that associates to phosphatidic acid under UASz control)

605581

w*;;  M{UASz-PASS-sfGFP }ZH-86Fb

3rd

22

PASS-StFP

(PA lipid biosensor.  mScarlet-i (StFP) fused to PASS domain that associates to phosphatidic acid under UASz control)

605582

w*;;  M{UASz-PASS-StFP }ZH-86Fb

3rd

22

Arp2-sfGFP

(superfolder GFP fused C-terminal to full-length Arp2 and expressed under control of the Arp2 promoter).

605605

w*  P{w+, Arp2-sfGFP}11

X

 

605606

w*; P{w+, Arp2-sfGFP}20

2nd

 

Arp3-StFP

(mScarlet-i (StFP) fused C-terminal to full-length Arp3 and expressed under control of the Arp3 promoter).

605607

w*  P{w+, Arp3-StFP}10

X

 

605608

w*; P{w+, Arp3-StFP}20

2nd

 

605609

w*; P{w+, Arp3-StFP}30

3rd

 

Wash

[GFP fusion protein under control of the UASp promoter (inducible)]

81639

w*; P{w[+mC]=UASp-GFP-Wash}15

X

 

81640

w*; P{w[+mC]=UASp-GFP-Wash}26

2nd

 

81641

w*; P{w[+mC]=UASp-GFP-Wash}35

3rd

 

Wash

[CherryFP fusion protein under control of the UASp promoter (inducible)]

81642

w*; P{w[+mC]=UASp-ChFP-Wash}10

2nd

18

81643

w*; P{w[+mC]=UASp-ChFP-Wash}31

3rd

18

Wash

[super folder GFP fusion protein under the control of the endogenous Wash promoter]

81644

w*; P{w[+mC]=Wash-sfGFP-Wash}104

2nd

14

SCAR

[GFP-SCAR fusion protein under control of endogenous SCAR promoter]

94927

w*;  P{GFP- SCAR }20; Dr/TM3,Sb

2nd

19

94928

w*;  Sp/CyO; P{GFP- SCAR }30

3rd

19

Rho1 GTPase 

(GFP fusion protein under control of endogenous Rho1 promoter)

9527

P{Rho1.GFP}, w*

X

5

9528

w*; P{Rho1.GFP}       

2nd

5

24762

w*; P{Rho1.GFP}30

3rd

5

Rho1 GTPase 

(ChFP fusion protein under control of endogenous Rho1 promoter)

52280

P{w[+mC]=mChFP-Rho1}10, w[*]

X

13

52281

w[*]; P{w[+mC]=mChFP-Rho1}21     

2nd

13

52282

w[*]; sna[Sco]/CyO; P{w[+mC]=mChFP-Rho1}31

3rd

13

Cdc42 GTPase 

(ChFP fusion protein under the control of the Sqh promoter)

42236

w*;  P{w[+mC]=sqh-ChFP-Cdc42}23

2nd

11

42237

w*;  P{w[+mC]=sqh-ChFP-Cdc42}3

3rd

11

Rac1 GTPase 

(GFP fusion protein under control of endogenous Rac1 promoter)

52283  

P{w[+mC]=GFP-Rac1}10, w[*]

X

13

52284

w[*]; P{w[+mC]=GFP-Rac1}20

2nd

13

52285

w[*]; P{w[+mC]=GFP-Rac1}30

3rd

13

Rac1 GTPase 

(CherryFP fusion protein under control of endogenous Rac1 promoter)

76266

w[*];  P{w[+mC]=ChFP-Rac1}21

2nd

14

76267

w[*];  P{w[+mC]=ChFP-Rac1}30

3rd

14

Rac2 GTPase 

(GFP fusion protein under control of endogenous Rac2 promoter)

52286

w[*]; P{w[+mC]=GFP-Rac2}21

2nd

13

52287

w[*]; P{w[+mC]=GFP-Rac2}31

3rd

13

Tumbleweed

(super folder GFP fusion protein under the control of the Sqh promoter)

76264

w*;  P{w[+mC]=sqh-sfGFP-Tum}20

2nd

14

76265

w*;  P{w[+mC]=sqh-sfGFP-Tum}30

3rd

14

Pebble

(GFP fusion protein under the control of the Sqh promoter)

76257

w* P{w[+mC]=sqh-Pbl-eGFP}10

X

14

76258

w*;  P{w[+mC]=sqh-Pbl-eGFP}30

3rd

14

RhoGEF2 

(super folder GFP fusion protein under the control of the Sqh promoter)

76259

w* P{w[+mC]=sqh-sfGFP-RhoGEF2}10

X

14

76260

w*;  P{w[+mC]=sqh-sfGFP-RhoGEF2}30

3rd

14

RhoGEF3  

(CherryFP fusion protein under control of the UASp promoter (inducible))

76261

w*; P{w[+mC]=UASp-ChFP-RhoGEF3}30

3rd

14

RhoGEF3

(super folder GFP fusion protein under control of the UASp promoter (inducible))

76262

w*; P{w[+mC]=UASp-sfGFP-RhoGEF3}20

2nd

14

76263

w*; P{w[+mC]=UASp-sfGFP-RhoGEF3}30

3rd

14

RhoGEF2 RNAi

(21nt shRNA from RhoGEF2 cloned into WALLIUM22; 99% knockdown by qPCR)

76255

M{w[+mC]=UAS-RhoGEF2.shRNA}ZH-86Fb

3rd

14

RhoGEF3 RNAi

(21nt shRNA from RhoGEF3 cloned into WALLIUM22; 92% knockdown by qPCR)

76256

M{w[+mC]=UAS-RhoGEF3.shRNA}ZH-86Fb

3rd

14

Rok

(GFP fusion protein under control of Sqh promoter)

52288

P{w[+mC]=sqh-GFP-rok}10, w[*]

X

13

52289

w[*]; P{w[+mC]=sqh-GFP-rok}30

3rd

13

Pak1

(GFP fusion protein under control of the UASp promoter (inducible)

52299

w[*]; P{w[+mC]=UASp-GFP-Pak1}300

3rd

13

Sticky

[super folder GFP fusion protein under the control of the Sqh promoter (constitutive)]

81645

w*; P{w[+mC]=sqh-sfGFP-Sti}10

X

 

81646

w*; P{w[+mC]=sqh-sfGFP-Sti}20

2nd

 

81647

w*; P{w[+mC]=sqh-sfGFP-Sti}32

3rd

 

Sticky

[CherryFP fusion protein under control of the UASp promoter (inducible)]

81648

w*; P{w[+mC]=UASp-ChFP-Sti}20/CyO

2nd

 

81649

w*; P{w[+mC]=UASp-ChFP-Sti}30/TM3

3rd

 

Rho family GTPase activity biosensors

(reporters for active Rho1, Rac, and/or Cdc42).

RokRBD

GFP fusion to Rok’s Rho1/Rac binding domain (RBD) expressed under control of the UASp promoter (inducible).

52290

w[*]; P{w[+mC]=UASp-rok.RBD-GFP}30

3rd

13

DiaRBD

GFP fusion to Diaphanous’s Rho1 binding domain (RBD) expressed under control of the UASp promoter (inducible).

52291

w[*]; P{w[+mC]=UASp-dia.RBD-GFP}20

2nd

13

52292

w[*]; P{w[+mC]=UASp-dia.RBD-GFP}37

3rd

13

CapuRBD

GFP fusion to Cappuccino’s Rho1 binding domain (RBD) expressed under control of the UASp promoter (inducible).

52293

w[*]; P{w[+mC]=UASp-capu.RBD-GFP}26

2nd

13

52294

w[*]; P{w[+mC]=UASp-capu.RBD-GFP}34

3rd

13

WashRBD

GFP fusion to Wash’s Rho1 binding domain (RBD) expressed under control of the UASp promoter (inducible).

52295

w[*]; P{w[+mC]=UASp-wash.RBD-GFP}25

2nd

13

52296

w[*]; P{w[+mC]=UASp-wash.RBD-GFP}34

3rd

13

PknRBD

GFP fusion to Pkn’s Rho/Rac binding domain (RBD) expressed under control of the Sqh promoter (constitutive).

52297

w[*]; P{w[+mC]=sqh-Pkn.RBD.G58A-eGFP}212a
and P{sqh-Pkn.RBD.G58A-eGFP}212b

2nd

13

52298

w[*]; P{w[+mC]=sqh-Pkn.RBD.G58A-eGFP}312a
and P{sqh-Pkn.RBD.G58A-eGFP}312b

3rd

13

WASpRBD

GFP fusion to WASp’s Cdc42 binding domain (RBD) expressed under control of the Sqh promoter (constitutive).

56745

w[*]; P{w[+mC]=sqh-WASp.RBD-GFP}278a
P{sqh-WASp.RBD-GFP}278b/CyO

2nd

13

56746

w[*]; P{w[+mC]=sqh-WASp.RBD-GFP}378a
P{sqh-WASp.RBD-GFP}378b

3rd

13

Pak1RBD

GFP fusion to Pak1’s Rac/Cdc42 binding domain (RBD) expressed under control of the Sqh promoter (constitutive) or under the control of the UASp promoter (conditional).

56549

w[*]; P{w[+mC]=sqh-Pak1.RBD-GFP}21

2nd

13

56550

w[*]; P{w[+mC]=sqh-Pak1.RBD-GFP}31/TM3, Sb[1]

3rd

13

56548

w[*]; P{w[+mC]=UASp-Pak.RBD-GFP}30/TM3, Sb[1]

3rd

13

Pak3RBD

GFP fusion to Pak3’s Rac/Cdc42 binding domain (RBD) expressed under control of the Sqh promoter (constitutive).

52303

w[*]; P{w[+mC]=sqh-Pak3.RBD-GFP}20

2nd

13

52304

w[*]; P{w[+mC]=sqh-Pak3.RBD-GFP}30

3rd

13

Cappuccino

(inducible GFP fusion protein)

24763

P{UASp-GFP.Capu}10, w*

X

5

24764

w*; P{UASp-GFP.Capu}20

2nd

5

Spire – C isoform

(inducible GFP fusion protein)

24765

w*; P{UASp-GFP.SpireC}21

2nd

5

24766

w*; P{UASp-GFP.SpireC}32

3rd

5

Spire – D isoform

(inducible GFP fusion protein)

24767

w*; P{UASp-GFP.SpireD}30

3rd

5

sisyphus, unconventional myosin 10A (myosin XV homolog)

(inducible RFP or GFP fusion proteins)

24780

P{UASp-Myo10A.GFP}10, w*

X

7

24781

w*; P{UASp-Myo10A.GFP}20

2nd

7

24782

w*; P{UASp-Myo10A.GFP}30        

3rd

7

24783

P{UASp-Myo10A.mRFP}11, w* 

X

7

24784

w*; P{UASp-Myo10A.mRFP}22     

2nd

7

24785

w*; P{UASp-Myo10A.mRFP}33 

3rd

7


References Cited

  1. Parkhurst SM and Ish-Horowicz D (1991). wimp, a dominant maternal-effect mutation, reduces transcription of a specific subset of segmentation genes in Drosophila.  Genes Dev5, 341-357.
  2. Magie CR, Meyer M, Gorsuch M and Parkhurst SM (1999). Mutations in the Rho1 small GTPase disrupt morphogenesis and segmentation during early Drosophila development.  Development 126, 5353-5364.
  3. Magie CR, Pinto-Santini D and Parkhurst SM (2002). Rho1 interacts with p120ctn and alpha-catenin, and regulates cadherin-based adherens junction formation during Drosophila development.  Development 129, 3771-3782.
  4. Rosenberg MI and Parkhurst SM (2002). Drosophila Sir2 is required for heterochromatic silencing and function of euchromatic Hairy/E(spl) family bHLH repressors in segmentation and sex determination.  Cell 109, 447-458.
  5. Rosales-Nieves AE, Johndrow JE, Keller LC, Magie CR, Pinto-Santini D and Parkhurst SM (2006). Coordination of microtubule and microfilament dynamics by Drosophila Rho1, Spire, and Cappuccino.  Nature Cell Biol8, 367-376.
  6. Linardopoulou EV, Parghi SS, Friedman C, Osborn GE, Parkhurst SM and Trask BJ (2007). Human subtelomeric WASH genes encode a new subclass of the WASP family. PLoS Genetics 3, e237.
  7. Liu R, Woolner S, Johndrow JE, Metzger D, Flores A and Parkhurst SM (2008). Sisyphus, the Drosophila myosin XV homolog, traffics within filopodia transporting key sensory and adhesion cargos. Development 135, 53-63.
  8. Liu R, Abreu-Blanco MT, Barry KC, Linardopoulou EV, Osborn GE and Parkhurst SM (2009). Wash functions downstream of Rho and links linear and branched action nucleation factors.  Development 136, 2849-2860.
  9. Barry KC, Abed M, Kenyagin D, Werwie TR, Boico O, Orian A and Parkhurst SM (2011). The Drosophila STUbL protein Degringolade limits HES function during embryogenesis. Development 138, 1759-1769.
  10. Abreu-Blanco MT, Verboon JM and Parkhurst SM (2011). Cell wound repair in Drosophila occurs through three distinct phases of membrane and cytoskeletal remodeling. J. Cell Biol193, 455-464.
  11. Abreu-Blanco MT, Verboon JM, Liu R, Watts JJ and Parkhurst SM (2012). Drosophila embryos close epithelial wounds using a combination of cellular protrusions and an actomyosin purse string. J. Cell Sci., 125, 5984-5997. 
  12. Rodriguez-Mesa E, Abreu-Blanco MT, Rosales-Nieves AE and Parkhurst SM (2012). Developmental expression of Drosophila Wiskott Aldrich Syndrome family proteins. Dev. Dyn241, 608-626
  13. Abreu-Blanco MT, Verboon JM and Parkhurst SM (2014). Coordination of Rho family GTPase activities to orchestrate cytoskeleton responses during cell wound repair. Curr. Biol., 24: 144-155.
  14. Verboon JM, Decker JR, Nakamura M and Parkhurst SM. Wash exhibits context dependent phenotypes and, along with the WASH Regulatory Complex, regulates Drosophila oogenesis. J. Cell Sci. 131(8), pii: jcs211573. doi: 10.1242/jcs.211573 (2018).   Erratum in: J. Cell Sci. 131(9), pii: jcs219212. doi: 10.1242/jcs.219212 (2018)
  15. Verboon JM, Mahmut D, Kim AR, Nakamura M, Abdulhay NJ, Nandakumar SK, Gupta N, Akie TE, Geddis AE, Manes B, Kapp ME, Hofmann I, Gabriel SB, Klein DE, Williams DA, Frangoul HA, Parkhurst SM, Crane GM, Cantor AB and Sankaran VG. (2020). Infantile Myelofibrosis and Myeloproliferation with CDC42 Dysfunction. J. Clin. Immunol. 40(4):554-566.  https://doi.org/10.1007/s10875-020-00778-7.
  16. Nakamura M, Verboon JM, Prentiss CL and Parkhurst SM (2020). The kinesin-like protein Pavarotti functions non-canonically to regulate actin dynamics. J. Cell Biol. 219(9):e201912117.  doi: 10.1083/jcb.201912117. PMID: 32673395.
  17. Nakamura M, Verboon JM, Allen TE, Abreu-Blanco MT, Liu R, Dominguez AN, Delrow JJ and Parkhurst SM (2020). Autocrine insulin pathway signaling regulates actin dynamics in cell wound repair. PLoS Genetics 16(12): e1009186. https://doi.org/10.1371/journal.pgen.1009186 
  18. Verboon JM, Rahe TK, Rodriguez-Mesa E and Parkhurst SM (2015). Wash functions downstream of Rho1 GTPase in a subset of Drosophila immune cell developmental migrations. Mol. Biol. Cell 26(9),1665-1674. doi:10.1091/mbc.E14-08-1266
  19. Hui J, Nakamura M, Dubrulle J and Parkhurst SM (2022). Coordinated efforts of different actin filament populations are needed for optimal cell wound repair. Mol. Biol. Cell 34(3):ar15. doi: 10.1091/mbc.E22-05-0155.
  20. Davidson KA, Nakamura M, Verboon JM and Parkhurst SM (2023). The Centralspindlin proteins Pavarotti and Tumbleweed function in Nuclear Envelope budding. J. Cell Biol. 222 (8): e202211074.  https://doi.org/10.1083/jcb202211074
  21. Stjepić V*, Nakamura M*, Hui J* and Parkhurst SM. (2024). Two Septin Complexes Mediate Actin Dynamics during Cell Wound Repair. Cell Reports 43: 114215.  https://doi.org/10.1016/j.celrep.2024.114215.
  22. Nakamura M and Parkhurst SM. (2024). Calcium influx rapidly establishes distinct spatial recruitments of Annexins to cell wounds. Genetics, iyae101, https://doi.org/10.1093/genetics/iyae101.