TY - JOUR

T1 - Interactome network analysis identifies multiple caspase-6 interactors involved in the pathogenesis of HD

AU - Riechers, Sean-Patrick

AU - Butland, Stefanie

AU - Deng, Yu

AU - Skotte, Niels

AU - Ehrnhoefer, Dagmar E

AU - Russ, Jenny

AU - Laine, Jean

AU - Laroche, Melissa

AU - Pouladi, Mahmoud A

AU - Wanker, Erich E

AU - Hayden, Michael R

AU - Graham, Rona K

N1 - © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

PY - 2016/4/15

Y1 - 2016/4/15

N2 - Caspase-6 (CASP6) has emerged as an important player in Huntington disease (HD), Alzheimer disease (AD) and cerebral ischemia, where it is activated early in the disease process. CASP6 also plays a key role in axonal degeneration, further underscoring the importance of this protease in neurodegenerative pathways. As a protein's function is modulated by its protein-protein interactions, we performed a high-throughput yeast-2-hybrid (Y2H) screen against ∼17,000 human proteins to gain further insight into the function of CASP6. We identified a high-confidence list of 87 potential CASP6 interactors. From this list, 61% are predicted to contain a CASP6 recognition site. Of nine candidate substrates assessed, six are cleaved by CASP6. Proteins that did not contain a predicted CASP6 recognition site were assessed using a LUMIER assay approach, and 51% were further validated as interactors by this method. Of note, 54% of the high-confidence interactors identified show alterations in human HD brain at the mRNA level, and there is a significant enrichment for previously validated huntingtin (HTT) interactors. One protein of interest, STK3, a pro-apoptotic kinase, was validated biochemically to be a CASP6 substrate. Furthermore, our results demonstrate that in striatal cells expressing mutant huntingtin (mHTT), an increase in full length and fragment levels of STK3 are observed. We further show that caspase-3 is not essential for the endogenous cleavage of STK3. Characterization of the interaction network provides important new information regarding key pathways of interactors of CASP6 and highlights potential novel therapeutic targets for HD, AD and cerebral ischemia.

AB - Caspase-6 (CASP6) has emerged as an important player in Huntington disease (HD), Alzheimer disease (AD) and cerebral ischemia, where it is activated early in the disease process. CASP6 also plays a key role in axonal degeneration, further underscoring the importance of this protease in neurodegenerative pathways. As a protein's function is modulated by its protein-protein interactions, we performed a high-throughput yeast-2-hybrid (Y2H) screen against ∼17,000 human proteins to gain further insight into the function of CASP6. We identified a high-confidence list of 87 potential CASP6 interactors. From this list, 61% are predicted to contain a CASP6 recognition site. Of nine candidate substrates assessed, six are cleaved by CASP6. Proteins that did not contain a predicted CASP6 recognition site were assessed using a LUMIER assay approach, and 51% were further validated as interactors by this method. Of note, 54% of the high-confidence interactors identified show alterations in human HD brain at the mRNA level, and there is a significant enrichment for previously validated huntingtin (HTT) interactors. One protein of interest, STK3, a pro-apoptotic kinase, was validated biochemically to be a CASP6 substrate. Furthermore, our results demonstrate that in striatal cells expressing mutant huntingtin (mHTT), an increase in full length and fragment levels of STK3 are observed. We further show that caspase-3 is not essential for the endogenous cleavage of STK3. Characterization of the interaction network provides important new information regarding key pathways of interactors of CASP6 and highlights potential novel therapeutic targets for HD, AD and cerebral ischemia.

KW - Binding Sites

KW - Caspase 6

KW - Cell Line

KW - Gene Expression Regulation

KW - Humans

KW - Huntingtin Protein

KW - Huntington Disease

KW - Models, Biological

KW - Protein Interaction Maps

KW - Protein Processing, Post-Translational

KW - Protein-Serine-Threonine Kinases

KW - Two-Hybrid System Techniques

KW - Journal Article

KW - Research Support, Non-U.S. Gov't

U2 - 10.1093/hmg/ddw036

DO - 10.1093/hmg/ddw036

M3 - Journal article

C2 - 26908611

VL - 25

SP - 1600

EP - 1618

JO - Human Molecular Genetics

JF - Human Molecular Genetics

SN - 0964-6906

IS - 8

ER -