Identification and characterization of a retina-specific calpain (Rt88) from rat

M. Azuma, C. Fukiage, M. Higashine, T. Nakajima, Hong Ma, Thomas (Tom) Shearer

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Purpose. To identify and characterize a newly discovered calpain termed Rt88 from rat retina. Methods. Rt88 in retina under normal physiological conditions was characterized in Sprague-Dawley rats of various ages by competitive RT-PCR, Northern blot analysis, cDNA cloning and sequencing. Recombinant Rt88 was expressed in the baculovirus system and characterized by casein zymography and immunoblotting. Results. Rt88 was sequenced and found to be similar to muscle calpain p94 except for three differences. A different exon 1 (as in lens Lp82 calpain) was present, and exons 15 and 16 in the unique IS2 region of muscle p94 were deleted. Of eleven tissues studied, mRNA for Rt88 was found only in retina where Rt88 increased with maturation and then remained constant. Casein zymography showed that rRt88 was proteolytically active after activation by calcium, but intact rRt88 was rapidly broken due to the presence of the IS1 region in domain II. Conclusions. Rt88 is a retina-specific, calcium activated protease from the calpain superfamily (EC 3.4.22.17) of cysteine proteases. Rt88 is a recently identified member of the AX1 subfamily of calpains showing alternative exon 1 usage. So far, all AX1 subfamily members are from eye. Rt88 may perform specific proteolytic functions during development, normal turnover, or pathological degeneration of retinal proteins.

Original languageEnglish (US)
Pages (from-to)710-720
Number of pages11
JournalCurrent Eye Research
Volume21
Issue number3
DOIs
StatePublished - 2000

Fingerprint

Calpain
Retina
Exons
Caseins
Muscles
Retinal Degeneration
Cysteine Proteases
Baculoviridae
Immunoblotting
Northern Blotting
Lenses
Sprague Dawley Rats
Organism Cloning
Complementary DNA
Calcium
Polymerase Chain Reaction
Messenger RNA
Proteins

Keywords

  • Calpain p94 isoform
  • Molecular biology
  • Proteinases
  • Rat
  • Retina

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems

Cite this

Identification and characterization of a retina-specific calpain (Rt88) from rat. / Azuma, M.; Fukiage, C.; Higashine, M.; Nakajima, T.; Ma, Hong; Shearer, Thomas (Tom).

In: Current Eye Research, Vol. 21, No. 3, 2000, p. 710-720.

Research output: Contribution to journalArticle

Azuma, M. ; Fukiage, C. ; Higashine, M. ; Nakajima, T. ; Ma, Hong ; Shearer, Thomas (Tom). / Identification and characterization of a retina-specific calpain (Rt88) from rat. In: Current Eye Research. 2000 ; Vol. 21, No. 3. pp. 710-720.
@article{bc279820a51e4ea88d2ecc0aa2fd6b8d,
title = "Identification and characterization of a retina-specific calpain (Rt88) from rat",
abstract = "Purpose. To identify and characterize a newly discovered calpain termed Rt88 from rat retina. Methods. Rt88 in retina under normal physiological conditions was characterized in Sprague-Dawley rats of various ages by competitive RT-PCR, Northern blot analysis, cDNA cloning and sequencing. Recombinant Rt88 was expressed in the baculovirus system and characterized by casein zymography and immunoblotting. Results. Rt88 was sequenced and found to be similar to muscle calpain p94 except for three differences. A different exon 1 (as in lens Lp82 calpain) was present, and exons 15 and 16 in the unique IS2 region of muscle p94 were deleted. Of eleven tissues studied, mRNA for Rt88 was found only in retina where Rt88 increased with maturation and then remained constant. Casein zymography showed that rRt88 was proteolytically active after activation by calcium, but intact rRt88 was rapidly broken due to the presence of the IS1 region in domain II. Conclusions. Rt88 is a retina-specific, calcium activated protease from the calpain superfamily (EC 3.4.22.17) of cysteine proteases. Rt88 is a recently identified member of the AX1 subfamily of calpains showing alternative exon 1 usage. So far, all AX1 subfamily members are from eye. Rt88 may perform specific proteolytic functions during development, normal turnover, or pathological degeneration of retinal proteins.",
keywords = "Calpain p94 isoform, Molecular biology, Proteinases, Rat, Retina",
author = "M. Azuma and C. Fukiage and M. Higashine and T. Nakajima and Hong Ma and Shearer, {Thomas (Tom)}",
year = "2000",
doi = "10.1076/0271-3683(200009)21:3;1-R;FT710",
language = "English (US)",
volume = "21",
pages = "710--720",
journal = "Current Eye Research",
issn = "0271-3683",
publisher = "Informa Healthcare",
number = "3",

}

TY - JOUR

T1 - Identification and characterization of a retina-specific calpain (Rt88) from rat

AU - Azuma, M.

AU - Fukiage, C.

AU - Higashine, M.

AU - Nakajima, T.

AU - Ma, Hong

AU - Shearer, Thomas (Tom)

PY - 2000

Y1 - 2000

N2 - Purpose. To identify and characterize a newly discovered calpain termed Rt88 from rat retina. Methods. Rt88 in retina under normal physiological conditions was characterized in Sprague-Dawley rats of various ages by competitive RT-PCR, Northern blot analysis, cDNA cloning and sequencing. Recombinant Rt88 was expressed in the baculovirus system and characterized by casein zymography and immunoblotting. Results. Rt88 was sequenced and found to be similar to muscle calpain p94 except for three differences. A different exon 1 (as in lens Lp82 calpain) was present, and exons 15 and 16 in the unique IS2 region of muscle p94 were deleted. Of eleven tissues studied, mRNA for Rt88 was found only in retina where Rt88 increased with maturation and then remained constant. Casein zymography showed that rRt88 was proteolytically active after activation by calcium, but intact rRt88 was rapidly broken due to the presence of the IS1 region in domain II. Conclusions. Rt88 is a retina-specific, calcium activated protease from the calpain superfamily (EC 3.4.22.17) of cysteine proteases. Rt88 is a recently identified member of the AX1 subfamily of calpains showing alternative exon 1 usage. So far, all AX1 subfamily members are from eye. Rt88 may perform specific proteolytic functions during development, normal turnover, or pathological degeneration of retinal proteins.

AB - Purpose. To identify and characterize a newly discovered calpain termed Rt88 from rat retina. Methods. Rt88 in retina under normal physiological conditions was characterized in Sprague-Dawley rats of various ages by competitive RT-PCR, Northern blot analysis, cDNA cloning and sequencing. Recombinant Rt88 was expressed in the baculovirus system and characterized by casein zymography and immunoblotting. Results. Rt88 was sequenced and found to be similar to muscle calpain p94 except for three differences. A different exon 1 (as in lens Lp82 calpain) was present, and exons 15 and 16 in the unique IS2 region of muscle p94 were deleted. Of eleven tissues studied, mRNA for Rt88 was found only in retina where Rt88 increased with maturation and then remained constant. Casein zymography showed that rRt88 was proteolytically active after activation by calcium, but intact rRt88 was rapidly broken due to the presence of the IS1 region in domain II. Conclusions. Rt88 is a retina-specific, calcium activated protease from the calpain superfamily (EC 3.4.22.17) of cysteine proteases. Rt88 is a recently identified member of the AX1 subfamily of calpains showing alternative exon 1 usage. So far, all AX1 subfamily members are from eye. Rt88 may perform specific proteolytic functions during development, normal turnover, or pathological degeneration of retinal proteins.

KW - Calpain p94 isoform

KW - Molecular biology

KW - Proteinases

KW - Rat

KW - Retina

UR - http://www.scopus.com/inward/record.url?scp=0034536146&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034536146&partnerID=8YFLogxK

U2 - 10.1076/0271-3683(200009)21:3;1-R;FT710

DO - 10.1076/0271-3683(200009)21:3;1-R;FT710

M3 - Article

C2 - 11120559

AN - SCOPUS:0034536146

VL - 21

SP - 710

EP - 720

JO - Current Eye Research

JF - Current Eye Research

SN - 0271-3683

IS - 3

ER -