Polymerase delta-interacting protein 38 (PDIP38) modulates the stability and activity of the mitochondrial AAA+ protease CLPXP

Philip R Strack,Matthew A Perugini,Liz J Valente,David A Dougan,Lauren M Angley,Erica J Brodie,Tamanna Saiyed,Kornelius Zeth,Kaye N Truscott,Bradley R Lowth,Hanmiao Zhan,Verena J Schuenemann

doi:10.1038/s42003-020-01358-6

Abstract

Over a decade ago Polymerase δ interacting protein of 38 kDa (PDIP38) was proposed to play a role in DNA repair. Since this time, both the physiological function and subcellular location of PDIP38 has remained ambiguous and our present understanding of PDIP38 function has been hampered by a lack of detailed biochemical and structural studies. Here we show, that human PDIP38 is directed to the mitochondrion in a membrane potential dependent manner, where it resides in the matrix compartment, together with its partner protein CLPX. Our structural analysis revealed that PDIP38 is composed of two conserved domains separated by an α/β linker region. The N-terminal (YccV-like) domain of PDIP38 forms an SH3-like β-barrel, which interacts specifically with CLPX, via the adaptor docking loop within the N-terminal Zinc binding domain of CLPX. In contrast, the C-terminal (DUF525) domain forms an immunoglobin-like β-sandwich fold, which contains a highly conserved putative substrate binding pocket. Importantly, PDIP38 modulates the substrate specificity of CLPX and protects CLPX from LONM-mediated degradation, which stabilises the cellular levels of CLPX. Collectively, our findings shed new light on the mechanism and function of mitochondrial PDIP38, demonstrating that PDIP38 is a bona fide adaptor protein for the mitochondrial protease, CLPXP.

Highlights

Over a decade ago Polymerase δ interacting protein of 38 kDa (PDIP38) was proposed to play a role in DNA repair
Human PDIP38 is targeted to the mitochondrial matrix, where it associates with CLPX
PDIP38 was originally described as a nuclear protein, it has been reported to localise to several subcellular compartments, including the mitochondrion, the cytosol and even the plasma membrane

Summary

Introduction

Over a decade ago Polymerase δ interacting protein of 38 kDa (PDIP38) was proposed to play a role in DNA repair. PDIP38 has been identified in the nucleus[36,37], the mitochondrion[15,38], the cytoplasm[39,40,41] and the plasma membrane[39,40], where it is proposed to mediate a variety of cellular functions, including cell proliferation[40], regulation of the extracellular matrix[42], oxidative signalling and cell migration[43], Tau aggregation[44], cancer[45] and DNA repair[37,39,41,46] Several of these cellular studies have incorporated EGFP-tagged versions of PDIP38 (which when attached to the N terminus of PDIP38 prevent its targeting to the mitochondrion) and very few of the putative interactions have been validated in vitro. We propose that mitochondrial PDIP38 is a specialised adaptor protein for the CLPXP protease Consistent with this role, PDIP38 is composed of two domains, an N-terminal “YccV-like” domain, which docks to a specific adaptor binding loop within the zinc-binding domain (ZBD) of CLPX, and a C-terminal domain of unknown function (DUF525), which forms an immunoglobulin-like fold. We propose that mitochondrial PDIP38 represents the first adaptor protein for the AAA+ protease, CLPXP

Methods

Results

Conclusion