Reversion mutation of cDNA CA8-204 minigene construct produces a truncated functional peptide that regulates calcium release in vitro and produces profound analgesia in vivo

Udita Upadhyay; Gerald Z Zhuang; Luda Diatchenko; Marc Parisien; Yuan Kang; Konstantinos D Sarantopoulos; Eden R Martin; Shad B Smith; William Maixner; Roy C Levitt

doi:10.1007/s00335-020-09848-y

Reversion mutation of cDNA CA8-204 minigene construct produces a truncated functional peptide that regulates calcium release in vitro and produces profound analgesia in vivo

Mamm Genome. 2020 Dec;31(9-12):287-294. doi: 10.1007/s00335-020-09848-y. Epub 2020 Nov 28.

Authors

Udita Upadhyay¹, Gerald Z Zhuang¹, Luda Diatchenko², Marc Parisien², Yuan Kang¹, Konstantinos D Sarantopoulos^{1

3}, Eden R Martin^{4

5}, Shad B Smith⁶, William Maixner⁶, Roy C Levitt^{7

8

9

10

11}

Affiliations

¹ Department of Anesthesiology, Perioperative Medicine and Pain Management, University of Miami Miller School of Medicine, Miami, FL, USA.
² Department of Anesthesiology, Alan Edwards Centre for Research on Pain, McGill University, Montreal, QC, Canada.
³ Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, FL, USA.
⁴ John T. MacDonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, Miami, FL, USA.
⁵ John P. Hussman Institute for Human Genomics, University of Miami Miller School of Medicine, Miami, FL, USA.
⁶ Department of Anesthesiology, Center for Translational Pain Medicine, Duke University School of Medicine, Durham, NC, USA.
⁷ Department of Anesthesiology, Perioperative Medicine and Pain Management, University of Miami Miller School of Medicine, Miami, FL, USA. rlevitt@med.miami.edu.
⁸ Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, FL, USA. rlevitt@med.miami.edu.
⁹ John T. MacDonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, Miami, FL, USA. rlevitt@med.miami.edu.
¹⁰ John P. Hussman Institute for Human Genomics, University of Miami Miller School of Medicine, Miami, FL, USA. rlevitt@med.miami.edu.
¹¹ University of Miami Miller School of Medicine, Rosenstiel Medical Sciences Building - Room 8010A (R-371), Miami, FL, 33136, USA. rlevitt@med.miami.edu.

PMID: 33247772
DOI: 10.1007/s00335-020-09848-y

Abstract

Intracellular calcium is critical in orchestrating neuronal excitability and analgesia. Carbonic anhydrase-8 (CA8) regulates intracellular calcium signaling through allosteric inhibition of neuronal inositol trisphosphate receptor 1 (ITPR1) to produce profound analgesia. Recently, we reported the "G" allele at rs6471859 represents cis-eQTL regulating alternative splicing of a 1697 bp transcript (CA8-204^G) with a retained intron, alternative polyadenylation site and a new stop codon producing a functional 26 kDa peptide with an extended exon 3. In this study we show the reversion mutation (G to C) at rs6471859 within the CA8-204^G expression vector also produced a stable 1697 bp transcript (CA8-204^C) coding for a smaller peptide (~ 22 kDa) containing only the first three CA8 exons. Surprisingly, this peptide inhibited ITPR1 (pITPR1) activation, ITPR1-mediated calcium release in vitro; and produced profound analgesia in vivo. This is the first report showing CA8-204^C codes for a functional peptide sufficient to regulate calcium signaling and produce profound analgesia.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Adenosine Triphosphate / metabolism
Analgesia*
Animals
Biomarkers, Tumor / chemistry
Biomarkers, Tumor / genetics*
Calcium / metabolism*
DNA, Complementary*
Dependovirus / genetics
Gene Transfer Techniques
Genetic Vectors / genetics
Humans
Mice
Mutation*
Pain / etiology
Pain / metabolism
Peptides / genetics*
Transduction, Genetic

Substances

Biomarkers, Tumor
DNA, Complementary
Peptides
Adenosine Triphosphate
CA8 protein, human
Calcium

Grants and funding

R21 NS105880/NS/NINDS NIH HHS/United States