km23-1 was originally identified as a TGFß receptor-interacting protein that 
plays an important role in TGFß signaling. Moreover, km23-1 is actually part of 
an ancient superfamily of NTPase-regulatory proteins, widely represented in 
archaea and bacteria. To further elucidate the function of km23-1, we identified 
novel protein interacting partners for km23-1 by using tandem affinity 
purification (TAP) and tandem mass spectrometry (MS). Here we show that km23-1 
interacted with a class of proteins involved in actin-based cell motility and 
modulation of the actin cytoskeleton. We further showed that km23-1 modulates 
the formation of a highly organized stress fiber network. More significantly, we 
demonstrated that knockdown (KD) of km23-1 decreased RhoA activation in Mv1Lu 
epithelial cells. Finally, our results demonstrated for the first time that 
depletion of km23-1 inhibited cell migration of human colon carcinoma cells 
(HCCCs) in wound-healing assays. Overall, our findings demonstrate that km23-1 
regulates RhoA and motility-associated actin modulating proteins, suggesting 
that km23-1 may represent a novel target for anti-metastatic therapy.