TY - JOUR
T1 - Mitochondrial and redox modifications in Huntington Disease induced pluripotent stem cells rescued by CRISPR/Cas9 CAGs targeting
AU - Lopes, Carla
AU - Tang, Yang
AU - Anjo, Sandra I.
AU - Manadas, Bruno
AU - Onofre, Isabel
AU - de Almeida, Luís P.
AU - Daley, George Q.
AU - Schlaeger, Thorsten M.
AU - Rego, Ana Cristina Carvalho
N1 - Funding Information:
We thank Dr. Mónica Zuzarte for electron microscopy images and to Dr. Henrique Girão and Teresa Ribeiro-Rodrigues for NTA analysis and useful discussions. Funding. This study was supported by the ‘FLAD Life Science 2020’ prize, funded by ‘Fundação Luso-Americana para o Desenvolvimento’ (FLAD), Portugal. The work was also financed by the European Regional Development Fund (ERDF), through the Centro 2020 Regional Operational Programme under project CENTRO-01-0145-FEDER-000012- HealthyAging2020, and through COMPETE 2020-Operational Programme for Competitiveness and Internationalization and Portuguese national funds via FCT–Fundação para a Ciência e a Tecnologia, under projects POCI-01-0145-FEDER-007440, POCI-01-0145-FEDER-029621, POCI-01-0145-FEDER-016428 (ref.: SAICTPAC/0010/2015), POCI-01-0145-FEDER-30943 (ref.: PTDC/MEC-PSQ/30943/2017), PTDC/MED-NEU/27946/2017; by The National Mass Spectrometry Network (RNEM) under the contract POCI-01-0145-FEDER-402-022125 (ref. ROTEIRO/0028/2013), UID/NEU/04539/2019 and UIDB/04539/2020.
Publisher Copyright:
© Copyright © 2020 Lopes, Tang, Anjo, Manadas, Onofre, de Almeida, Daley, Schlaeger and Rego.
PY - 2020/9/22
Y1 - 2020/9/22
N2 - Mitochondrial deregulation has gained increasing support as a pathological mechanism in Huntington’s disease (HD), a genetic-based neurodegenerative disorder caused by CAG expansion in the HTT gene. In this study, we thoroughly investigated mitochondrial-based mechanisms in HD patient-derived iPSC (HD-iPSC) and differentiated neural stem cells (NSC) versus control cells, as well as in cells subjected to CRISPR/Cas9-CAG repeat deletion. We analyzed mitochondrial morphology, function and biogenesis, linked to exosomal release of mitochondrial components, glycolytic flux, ATP generation and cellular redox status. Mitochondria in HD cells exhibited round shape and fragmented morphology. Functionally, HD-iPSC and HD-NSC displayed lower mitochondrial respiration, exosomal release of cytochrome c, decreased ATP/ADP, reduced PGC-1α and complex III subunit expression and activity, and were highly dependent on glycolysis, supported by pyruvate dehydrogenase (PDH) inactivation. HD-iPSC and HD-NSC mitochondria showed ATP synthase reversal and increased calcium retention. Enhanced mitochondrial reactive oxygen species (ROS) were also observed in HD-iPSC and HD-NSC, along with decreased UCP2 mRNA levels. CRISPR/Cas9-CAG repeat deletion in HD-iPSC and derived HD-NSC ameliorated mitochondrial phenotypes. Data attests for intricate metabolic and mitochondrial dysfunction linked to transcriptional deregulation as early events in HD pathogenesis, which are alleviated following CAG deletion.
AB - Mitochondrial deregulation has gained increasing support as a pathological mechanism in Huntington’s disease (HD), a genetic-based neurodegenerative disorder caused by CAG expansion in the HTT gene. In this study, we thoroughly investigated mitochondrial-based mechanisms in HD patient-derived iPSC (HD-iPSC) and differentiated neural stem cells (NSC) versus control cells, as well as in cells subjected to CRISPR/Cas9-CAG repeat deletion. We analyzed mitochondrial morphology, function and biogenesis, linked to exosomal release of mitochondrial components, glycolytic flux, ATP generation and cellular redox status. Mitochondria in HD cells exhibited round shape and fragmented morphology. Functionally, HD-iPSC and HD-NSC displayed lower mitochondrial respiration, exosomal release of cytochrome c, decreased ATP/ADP, reduced PGC-1α and complex III subunit expression and activity, and were highly dependent on glycolysis, supported by pyruvate dehydrogenase (PDH) inactivation. HD-iPSC and HD-NSC mitochondria showed ATP synthase reversal and increased calcium retention. Enhanced mitochondrial reactive oxygen species (ROS) were also observed in HD-iPSC and HD-NSC, along with decreased UCP2 mRNA levels. CRISPR/Cas9-CAG repeat deletion in HD-iPSC and derived HD-NSC ameliorated mitochondrial phenotypes. Data attests for intricate metabolic and mitochondrial dysfunction linked to transcriptional deregulation as early events in HD pathogenesis, which are alleviated following CAG deletion.
KW - Huntington disease
KW - Induced pluripotent stem cells
KW - Mitochondrial dysfunction
KW - Neural stem cells
KW - Reactive oxygen species
KW - Transcriptional deregulation
UR - http://www.scopus.com/inward/record.url?scp=85092059116&partnerID=8YFLogxK
U2 - 10.3389/fcell.2020.576592
DO - 10.3389/fcell.2020.576592
M3 - Article
C2 - 33072759
AN - SCOPUS:85092059116
SN - 2296-634X
VL - 8
JO - Frontiers in Cell and Developmental Biology
JF - Frontiers in Cell and Developmental Biology
M1 - 576592
ER -