Increasing evidence indicated that the protein factors involved in DNA mismatch repair (MMR) possess meiotic functions beyond the scope of DNA mismatch correction. The important roles of MMR components in meiotic processes have been highlighted by the recent identification of two additional members of the mammalian MutS homologs, MSH4 and MSH5. Mammalian MSH4 and MSH5 proteins form a heterodimeric complex and play an important role in the meiotic processes. As a step forward to the understanding of the molecular mechanisms underlying the roles of these two mammalian MutS homologues, here we have identified von Hippel-Lindau (VHL) tumor suppressor-binding protein 1 (VBP1) as an interacting protein partner for human MSH4 (hMSH4). In addition, we have characterized a hMSH4 splicing variant (hMSH4sv) encoding a truncated form of hMSH4. The protein encoded by hMSH4sv was unable to interact with hMSH5, but it retained the capacity to interact with VBP1. It is conceivable that hMSH4 and hMSH4sv can carry out different but overlapping functions by differential protein interactions, and, therefore, hMSH4sv might represent a separation-of-function alternative form of the hMSH4 protein. hMSH4 and VBP1 proteins were colocalized in mammalian cells. Three-hybrid analysis suggested that VBP1 could compete with hMSH5 for the binding of hMSH4. Thus, hMSH4 may be involved in diverse cellular processes through interaction with different protein partners, and the levels of VBP1 protein expression in cells could potentially affect the availability of the hMSH4-hMSH5 hetero-complex.