We have previously described a novel modulator of the actin cytoskeleton that also regulates Ras and mitogen-activated protein kinase activities in TGFβ-sensitive epithelial cells. Here we examined the functional role of this signaling regulatory protein (km23-1) in mediating the migration, invasion, and tumor growth of human colorectal carcinoma (CRC) cells. We show that small interfering RNA (siRNA) depletion of km23-1 in human CRC cells inhibited constitutive extracellular signal-regulated kinase (ERK) activation, as well as pro-invasive ERK effector functions that include phosphorylation of Elk-1, constitutive regulation of c-Fos-DNA binding, TGFβ1 promoter transactivation, and TGFβ1 secretion. In addition, knockdown of km23-1 reduced the paracrine effects of CRC cell-secreted factors in conditioned medium and in fibroblast co-cultures. Moreover, km23-1 depletion in human CRC cells reduced cell migration and invasion, as well as expression of the ERK-regulated, metastasis-associated scaffold protein Ezrin. Finally, km23-1 inhibition significantly suppressed tumor formation in vivo. Thus, our results implicate km23-1 as a novel anti-metastasis target for human colon carcinoma cells, capable of decreasing tumor growth and invasion via a mechanism involving suppression of various pro-migratory features of CRC. These include a reduction in ERK signaling, diminished TGFβ1 production, decreased expression of the plasma membrane-cytoskeletal linker Ezrin, as well as attenuation of the paracrine effects of colon carcinoma-secreted factors on fibroblast migration and mitogenesis. As such, km23-1 inhibitors may represent a viable therapeutic strategy for interfering with colon cancer progression and invasion.