RAP80, a member of the BRCA1-A complex, is a well-known crucial regulator of cell cycle checkpoint and DNA damage repair in the nucleus. However, it is still unclear whether Rap80 localizes to another region outside the nucleus and plays different roles with its partners. Here, we found mitochondrial p32 as a novel binding partner of RAP80 by using yeast two-hybrid screening. RAP80 directly binds the internal region of p32 through its arginine rich C-terminal domain. Based on the interaction, we showed that a subset of RAP80 localizes to mitochondria where p32 exists. Loss of function study revealed that RAP80 deficiency reduces the protein level of p32 and p32 dependent mitochondrial translating proteins such as Rieske and COX1. As a result, mitochondrial membrane potential and oxygen consumption are reduced in RAP80 knockdown cells, indicating mitochondrial dysfunction. Our study identifies a novel interaction between RAP80 and p32, which is important for preserving intact mitochondrial function. (C) 2017 Elsevier Inc. All rights reserved.