Fig 2: ATHB17?113 forms homodimers and makes heterodimers with maize HD-Zip II proteins.Maize leaf protoplasts were (A) mock- transformed or transformed with constructs expressing CFP, MYC::HA and CFP tagged ATHB17?113 alone or CFP and ATHB17?113::CFP in combination with ATHB17?113::MYC construct, (B) transformed (filled bars) with constructs expressing CFP or each of the CFP tagged HD-Zip IIs alone or co-transformed (empty bars) with the construct expressing ATHB17?113::MYC-HA. Cellular extracts were co-immunoprecipitated using anti-MYC antibody and the immunoprecipitated complex was probed with anti-CFP antibody and visualized using phycoerythrin-conjugated reporter molecules. One HD-Zip I, Zmhdz3, was used to evaluate interaction with ATHB17?113. Nomenclature of HD-Zip II and HD-Zip I constructs is based on Zhao et al.[22].

Fig 3: LNX1 activates RhoC by inhibiting its interaction with RhoGDIa. (A) Western blot analysis of transfected HEK293T cells expressing DsRedM (control), Smurf1-MycFLAG or LNX1-MycFLAG proteins. Immunoblot with Myc shows overexpression of Smurf1 or LNX1. Immunoblot for RhoA shows reduction in RhoA upon Smurf1 overexpression. Immunoblot for RhoC shows that LNX1 overexpression does not affect endogenous RhoC protein level. H3 is a loading control. (B) Rhotekin Rho Binding Domain (RRBD) pull down from transfected HEK293T cells expressing DsRedM (control) or LNX1-MycFLAG protein. LNX1 overexpression results in significant increase in the amount of endogenous RhoC pulled down by RRBD reflecting increase in the amount of GTP-bound (active) form of RhoC. Graph quantifies results from n = 5 biologically independent experiments. **P = 0.0037. Data are presented as the mean ± s.d. (C) PLA in 293-CDC42 control or LNX1-overexpressing cells using HA/RhoGDIa or RhoC/RhoGDIa probes. DAPI (blue), PLA signal (orange). Scale bar, 10 µm. Graph quantifies results from three biologically independent experiments. Data are presented as individual data points in gray circles and their mean ± s.d. within each of 3 independent biological replicates as triangles color coded by experiment. (D) Filamentous actin content (phalloidin, white) of ReN cells transduced with a lentivirus to overexpress LNX1-V5 with or without RhoA or RhoC knock-down. Immunolabels with anti-eGFP, anti-tGFP or anti-V5 (orange) identify transduced cells. Scale bar, 10 µm. Graphs are presented as individual data points from all three biological replicates in gray circles and their mean ± s.d. as triangles color coded by experiment. All P values were obtained using a two-tailed paired t-test.