| Antibodies |
| Rabbit monoclonal phospho-Akt (Thr308) (D25E6) XP®
|
Cell Signaling Technology |
Cat #13038S; RRID: AB_2629447 |
| Rabbit monoclonal phospho-Akt (Ser473) (D9E) XP®
|
Cell Signaling Technology |
Cat #4060S; RRID: AB_2315049 |
| Rabbit polyclonal Akt |
Cell Signaling Technology |
Cat #9272S; RRID: AB_329827 |
| Mouse monoclonal p21 Ras |
Cytoskeleton Inc. |
Cat #AESA02 |
| Rabbit polyclonal GST |
EMD Millipore |
Cat # AB3282; RRID: AB_91439 |
| Goat anti-rabbit IRDye 680RD-conjugated secondary |
Li-Cor |
Cat #925-68071; RRID: AB_2721181 |
| Goat anti-mouse IRDye 800CW-conjugated secondary |
Li-Cor |
Cat #925-32210; RRID: AB_2687825 |
| Bacterial and virus strains |
| CIBN-CAAX expressing 3rd generation Lentivirus |
this paper |
N/A |
| LifeActmiRFP703 expressing 3rd generation Lentivirus |
this paper |
N/A |
|
Klebsiella aerogenes
|
Lab stock |
N/A |
| Chemicals, peptides, and recombinant proteins |
| RPMI medium 1640 |
Gibco |
Cat #22400-089 |
| Heat-inactivated fetal bovine serum |
Thermo Fisher Scientific |
Cat#16140071 |
| Penicillin-streptomycin |
Thermo Fisher Scientific |
Cat#15140122 |
| DMSO |
Sigma-Aldrich |
Cat # D2650 |
| DMEM medium |
Sigma-Aldrich |
Cat #D6429 |
| DMEM medium |
Gibco |
Cat #10569-010 |
| HL-5 medium |
Lab stock |
N/A |
| Fibronectin |
Sigma-Aldrich |
Cat #F4759-2MG |
| N-Formyl-Met-Leu-Phe |
Sigma-Aldrich |
Cat #47729 |
| FKP-(D-Cha)-Cha-r |
Anaspec |
Cat #65121 |
| Leukotriene B4 |
Cayman Chemical |
Cat #20110 |
| AS605240 |
Sigma-Aldrich |
Cat #A0233 |
| LY294002 |
Invitrogen |
Cat #PHZ1144 |
| PP242 |
EMD Millipore |
Cat #475988 |
| Rapamycin |
Cayman Chemical |
Cat #13346 |
| Pertussis toxin |
Sigma-Aldrich |
Cat #516560-50UG |
| Gallein |
Sigma-Aldrich |
Cat #371708-5MG |
| CK-666 |
EMD Millipore |
Cat #182515 |
| Janelia Fluor 646 HaloTag |
Promega Corporation |
Cat # GA1120 |
| Puromycin |
Sigma-Aldrich |
Cat # P8833 |
| Blasticidine S |
Sigma-Aldrich |
Cat #15205 |
| Hygromycin B |
Thermo Fisher Scientific |
Cat#10687010 |
| G418 sulphate |
Thermo Fisher Scientific |
Cat #10131035 |
| Lipofectamine 3000 |
Invitrogen |
Cat #L3000-008 |
| Polybrene |
Sigma-Aldrich |
Cat #TR1003 |
| Protease inhibitor cocktail |
Roche |
Cat#11836170001 |
| Pre-cast 4–15% polyacrylamide gel |
Bio-Rad |
Cat#5671085 |
| Intercept blocking buffer |
Li-Cor |
Cat #927-60001 |
| Critical commercial assays |
| Neon transfection kit |
Invitrogen |
Cat #MPK10025B |
| Amaxa cell line kit V |
Lonza |
Cat #VACA-1003 |
| Ras pull-down activation assay biochem kit |
Cytoskeleton, Inc. |
Cat #BK008S |
| Experimental models: Cell lines |
| Female human neutrophil-like HL-60 cell line |
Millius et al142,143
|
N/A |
| Male mouse monocyte/macrophage RAW 264.7 cell line |
Meshik et al144
|
N/A |
| Female human embryonic kidney HEK293T cell line |
Lab stock |
N/A |
|
Dictyostelium discoideum cells of the axenic strain AX2 |
Lab stock |
N/A |
| Oligonucleotides |
| Primers (P1-P26) for cloning, See Table S1
|
This paper |
N/A |
| Recombinant DNA |
| CIBN-CAAX/pLJM1 |
This paper |
Addgene Plasmid #201749 |
| LifeActmiRFP703/pLJM1 |
This paper |
Addgene Plasmid #201750 |
| SC0085 transposase expression plasmid |
Yusa et al145
|
N/A |
| pFUW2-RFP-PH-Akt |
Zhan et al146
|
N/A |
| pEGFPN1-Lifeact-7Alinker-Halo7 |
Shirai et al147
|
N/A |
| pMD2.G |
Didier Trono Lab (EPFL, Switzerland) (unpublished) |
Addgene Plasmid #12259 |
| pMDLg/pRRE |
Dull et al149
|
Addgene Plasmid #12251 |
| pRSV-Rev |
Dull et al149
|
Addgene Plasmid #12253 |
| CRY2PHR-mcherry2-CTRL/SC0030 |
This paper |
Addgene Plasmid #201751 |
| CRY2PHR-mcherry2-HRas G12V ΔCAAX/SC0030 |
This paper |
Addgene Plasmid #201752 |
| CRY2PHR-mcherry2-KRas4B G12V ΔCAAX/SC0030 |
This paper |
Addgene Plasmid #201753 |
| CRY2PHR-mcherry2-RasGRP4/SC0030 |
This paper |
Addgene Plasmid #201754 |
| CRY2PHR-mcherry2-RASAL3/SC0030 |
This paper |
Addgene Plasmid #201755 |
| CRY2PHR-mcherry2-Akt1/SC0030 |
This paper |
Addgene Plasmid #201756 |
| CRY2PHR-mcherry2-Akt1T308A/SC0030 |
This paper |
Addgene Plasmid #201757 |
| CRY2PHR-mcherry2-Akt2/SC0030 |
This paper |
Addgene Plasmid #201758 |
| pCIBN-CAAX |
Idevall-Hagren et al151
|
Addgene Plasmid #79574 |
| pCRY2PHR-mcherry2-RasGRP4 |
This paper |
Addgene Plasmid #201759 |
| pCRY2PHR-mcherry2-RASAL3 |
This paper |
Addgene Plasmid #201760 |
| N150-Venus-iLID/pDM358 |
This paper |
Addgene Plasmid #201763 |
| tgRFPt-SSPB R73Q-CTRL/pCV5 |
This paper |
Addgene Plasmid #201761 |
| tgRFPt-SSPB R73Q-PKBA/pCV5 |
This paper |
Addgene Plasmid #201762 |
| Software and algorithms |
| Fiji (ImageJ) Ver 1.52i |
N/A |
http://imagej.nih.gov/ij
|
| GraphPad Prism Ver 8.00 |
N/A |
https://graphpad.com
|
| MATLAB Ver R2019a |
N/A |
https://mathworks.com
|
| Other |
| Neon electroporation system |
Invitrogen |
Cat #MPK5000 |
| 0.1 cm-gap cuvette |
Bio-Rad |
Cat#1652089 |
| Gene Pulser Xcell Electroporation Systems |
Bio-Rad |
Cat#1652660 |
| Amaxa Nucleofector II device |
Lonza |
Cat #LOAAB-1001 |
| HPLS-45 470 nm LED controller system |
LightSpeed Technologies |
Cat #HPLS-R1000-3DExt |
| 10 cm cell culture dish |
Greiner Bio-One |
Cat#664160 |
| 6-well cell culture plate |
Greiner Bio-One |
Cat#657160 |
| 24-well cell culture plate |
Greiner Bio-One |
Cat#662160 |
| 48-well cell culture plate |
Sarstedt |
Cat #83.3923 |
| 8-well chambered coverglass |
LAB-TEK |
Cat#155409 |
| Chemotaxis μ-slides |
Ibidi |
Cat #80322 |
| PVDF membrane |
Bio-Rad |
Cat #162-0262 |
| Trans-Blot Turbo semi-dry transfer system |
Bio-Rad |
Cat #1704150EDU |
