Human melanocytes and melanomas express novel mRNA isoforms of the tyrosinase-related protein-2/DOPAchrome tautomerase gene: Molecular and functional characterization

Patrizia Pisarra, Raffaella Lupetti, Anna Palumbo, Alessandra Napolitano, Giuseppe Prota, Giorgio Parmiani, Andrea Anichini, Marialuisa Sensi

Research output: Contribution to journalArticle

Abstract

We previously reported that a melanoma antigen, recognized by tumor- specific cytotoxic T lymphocytes, was encoded by intron sequences retained in a partially spliced transcript of the tyrosinase-related protein-2/DOPAchrome tautomerase gene. At difference with the mRNA encoding tyrosinase-related protein-2, this anomalous transcript was not expressed in melanocytes. This study examined whether neoplastic and/or normal cells of the melanocytic lineage could express additional forms of tyrosinase-related protein-2 mRNA. Screening of a melanoma-derived cDNA library with a tyrosinase-related protein-2 probe allowed identification of two novel isoforms. The first, tyrosinase-related protein-2-long tail, corresponds to the dominant transcript detected on melanomas and melanocytes by northern blot analysis. Tyrosinase-related protein-2-long tail is identical to the tyrosinase-related protein-2-encoding published cDNA sequence except for an extended 3'- untranslated region and is originated by alternative polyadenylation. This novel 3'-untranslated region contains an alternatively spliced, tyrosinase- related protein-2 last exon in the second isoform (tyrosinase-related protein-2-8b). The protein encoded by tyrosinase-related protein-2-8b is identical to tyrosinase-related protein-2 in its first 460 amino acids but possesses a different carboxyl-terminus devoid of transmembrane domain. Tyrosinase-related protein-2-long tail exhibited DOPA-chrome tautomerase activity, when transiently transfected into COS-7 cells. On the contrary, no detectable activity was exhibited by tyrosinase-related protein-2-8b. Reverse transcription-polymerase chain reaction analysis indicated that tyrosinase- related protein-2-long tail and tyrosinase-related protein-2-8b are expressed by tyrosinase-related protein-2-positive melanomas and normal melanocytes. Moreover all cell lines positive for tyrosinase-related protein-2 isoforms expressed tyrosinase and, all but one, tyrosinase-related protein-1. These data show that the human tyrosinase-related protein-2/DOPAchrome tautomerase gene can yield different isoforms by alternative poly(A) site usage or by alternative splicing. The pattern of expression of these isoforms suggest that they might play a part in the normal pathway of melanin biosynthesis.

Original languageEnglish
Pages (from-to)48-56
Number of pages9
JournalJournal of Investigative Dermatology
Volume115
Issue number1
DOIs
Publication statusPublished - 2000

Fingerprint

RNA Isoforms
Melanocytes
Melanoma
Genes
Protein Isoforms
dopachrome isomerase
Tail
3' Untranslated Regions
Melanoma-Specific Antigens

Keywords

  • Alternative polyadenylation
  • Alternative splicing
  • Melanoma antigens
  • Pigmentation

ASJC Scopus subject areas

  • Dermatology

Cite this

Human melanocytes and melanomas express novel mRNA isoforms of the tyrosinase-related protein-2/DOPAchrome tautomerase gene : Molecular and functional characterization. / Pisarra, Patrizia; Lupetti, Raffaella; Palumbo, Anna; Napolitano, Alessandra; Prota, Giuseppe; Parmiani, Giorgio; Anichini, Andrea; Sensi, Marialuisa.

In: Journal of Investigative Dermatology, Vol. 115, No. 1, 2000, p. 48-56.

Research output: Contribution to journalArticle

Pisarra, Patrizia ; Lupetti, Raffaella ; Palumbo, Anna ; Napolitano, Alessandra ; Prota, Giuseppe ; Parmiani, Giorgio ; Anichini, Andrea ; Sensi, Marialuisa. / Human melanocytes and melanomas express novel mRNA isoforms of the tyrosinase-related protein-2/DOPAchrome tautomerase gene : Molecular and functional characterization. In: Journal of Investigative Dermatology. 2000 ; Vol. 115, No. 1. pp. 48-56.
@article{b82616d049054c43b29cd47eb452f1f0,
title = "Human melanocytes and melanomas express novel mRNA isoforms of the tyrosinase-related protein-2/DOPAchrome tautomerase gene: Molecular and functional characterization",
abstract = "We previously reported that a melanoma antigen, recognized by tumor- specific cytotoxic T lymphocytes, was encoded by intron sequences retained in a partially spliced transcript of the tyrosinase-related protein-2/DOPAchrome tautomerase gene. At difference with the mRNA encoding tyrosinase-related protein-2, this anomalous transcript was not expressed in melanocytes. This study examined whether neoplastic and/or normal cells of the melanocytic lineage could express additional forms of tyrosinase-related protein-2 mRNA. Screening of a melanoma-derived cDNA library with a tyrosinase-related protein-2 probe allowed identification of two novel isoforms. The first, tyrosinase-related protein-2-long tail, corresponds to the dominant transcript detected on melanomas and melanocytes by northern blot analysis. Tyrosinase-related protein-2-long tail is identical to the tyrosinase-related protein-2-encoding published cDNA sequence except for an extended 3'- untranslated region and is originated by alternative polyadenylation. This novel 3'-untranslated region contains an alternatively spliced, tyrosinase- related protein-2 last exon in the second isoform (tyrosinase-related protein-2-8b). The protein encoded by tyrosinase-related protein-2-8b is identical to tyrosinase-related protein-2 in its first 460 amino acids but possesses a different carboxyl-terminus devoid of transmembrane domain. Tyrosinase-related protein-2-long tail exhibited DOPA-chrome tautomerase activity, when transiently transfected into COS-7 cells. On the contrary, no detectable activity was exhibited by tyrosinase-related protein-2-8b. Reverse transcription-polymerase chain reaction analysis indicated that tyrosinase- related protein-2-long tail and tyrosinase-related protein-2-8b are expressed by tyrosinase-related protein-2-positive melanomas and normal melanocytes. Moreover all cell lines positive for tyrosinase-related protein-2 isoforms expressed tyrosinase and, all but one, tyrosinase-related protein-1. These data show that the human tyrosinase-related protein-2/DOPAchrome tautomerase gene can yield different isoforms by alternative poly(A) site usage or by alternative splicing. The pattern of expression of these isoforms suggest that they might play a part in the normal pathway of melanin biosynthesis.",
keywords = "Alternative polyadenylation, Alternative splicing, Melanoma antigens, Pigmentation",
author = "Patrizia Pisarra and Raffaella Lupetti and Anna Palumbo and Alessandra Napolitano and Giuseppe Prota and Giorgio Parmiani and Andrea Anichini and Marialuisa Sensi",
year = "2000",
doi = "10.1046/j.1523-1747.2000.00023.x",
language = "English",
volume = "115",
pages = "48--56",
journal = "Journal of Investigative Dermatology",
issn = "0022-202X",
publisher = "Nature Publishing Group",
number = "1",

}

TY - JOUR

T1 - Human melanocytes and melanomas express novel mRNA isoforms of the tyrosinase-related protein-2/DOPAchrome tautomerase gene

T2 - Molecular and functional characterization

AU - Pisarra, Patrizia

AU - Lupetti, Raffaella

AU - Palumbo, Anna

AU - Napolitano, Alessandra

AU - Prota, Giuseppe

AU - Parmiani, Giorgio

AU - Anichini, Andrea

AU - Sensi, Marialuisa

PY - 2000

Y1 - 2000

N2 - We previously reported that a melanoma antigen, recognized by tumor- specific cytotoxic T lymphocytes, was encoded by intron sequences retained in a partially spliced transcript of the tyrosinase-related protein-2/DOPAchrome tautomerase gene. At difference with the mRNA encoding tyrosinase-related protein-2, this anomalous transcript was not expressed in melanocytes. This study examined whether neoplastic and/or normal cells of the melanocytic lineage could express additional forms of tyrosinase-related protein-2 mRNA. Screening of a melanoma-derived cDNA library with a tyrosinase-related protein-2 probe allowed identification of two novel isoforms. The first, tyrosinase-related protein-2-long tail, corresponds to the dominant transcript detected on melanomas and melanocytes by northern blot analysis. Tyrosinase-related protein-2-long tail is identical to the tyrosinase-related protein-2-encoding published cDNA sequence except for an extended 3'- untranslated region and is originated by alternative polyadenylation. This novel 3'-untranslated region contains an alternatively spliced, tyrosinase- related protein-2 last exon in the second isoform (tyrosinase-related protein-2-8b). The protein encoded by tyrosinase-related protein-2-8b is identical to tyrosinase-related protein-2 in its first 460 amino acids but possesses a different carboxyl-terminus devoid of transmembrane domain. Tyrosinase-related protein-2-long tail exhibited DOPA-chrome tautomerase activity, when transiently transfected into COS-7 cells. On the contrary, no detectable activity was exhibited by tyrosinase-related protein-2-8b. Reverse transcription-polymerase chain reaction analysis indicated that tyrosinase- related protein-2-long tail and tyrosinase-related protein-2-8b are expressed by tyrosinase-related protein-2-positive melanomas and normal melanocytes. Moreover all cell lines positive for tyrosinase-related protein-2 isoforms expressed tyrosinase and, all but one, tyrosinase-related protein-1. These data show that the human tyrosinase-related protein-2/DOPAchrome tautomerase gene can yield different isoforms by alternative poly(A) site usage or by alternative splicing. The pattern of expression of these isoforms suggest that they might play a part in the normal pathway of melanin biosynthesis.

AB - We previously reported that a melanoma antigen, recognized by tumor- specific cytotoxic T lymphocytes, was encoded by intron sequences retained in a partially spliced transcript of the tyrosinase-related protein-2/DOPAchrome tautomerase gene. At difference with the mRNA encoding tyrosinase-related protein-2, this anomalous transcript was not expressed in melanocytes. This study examined whether neoplastic and/or normal cells of the melanocytic lineage could express additional forms of tyrosinase-related protein-2 mRNA. Screening of a melanoma-derived cDNA library with a tyrosinase-related protein-2 probe allowed identification of two novel isoforms. The first, tyrosinase-related protein-2-long tail, corresponds to the dominant transcript detected on melanomas and melanocytes by northern blot analysis. Tyrosinase-related protein-2-long tail is identical to the tyrosinase-related protein-2-encoding published cDNA sequence except for an extended 3'- untranslated region and is originated by alternative polyadenylation. This novel 3'-untranslated region contains an alternatively spliced, tyrosinase- related protein-2 last exon in the second isoform (tyrosinase-related protein-2-8b). The protein encoded by tyrosinase-related protein-2-8b is identical to tyrosinase-related protein-2 in its first 460 amino acids but possesses a different carboxyl-terminus devoid of transmembrane domain. Tyrosinase-related protein-2-long tail exhibited DOPA-chrome tautomerase activity, when transiently transfected into COS-7 cells. On the contrary, no detectable activity was exhibited by tyrosinase-related protein-2-8b. Reverse transcription-polymerase chain reaction analysis indicated that tyrosinase- related protein-2-long tail and tyrosinase-related protein-2-8b are expressed by tyrosinase-related protein-2-positive melanomas and normal melanocytes. Moreover all cell lines positive for tyrosinase-related protein-2 isoforms expressed tyrosinase and, all but one, tyrosinase-related protein-1. These data show that the human tyrosinase-related protein-2/DOPAchrome tautomerase gene can yield different isoforms by alternative poly(A) site usage or by alternative splicing. The pattern of expression of these isoforms suggest that they might play a part in the normal pathway of melanin biosynthesis.

KW - Alternative polyadenylation

KW - Alternative splicing

KW - Melanoma antigens

KW - Pigmentation

UR - http://www.scopus.com/inward/record.url?scp=0033939851&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033939851&partnerID=8YFLogxK

U2 - 10.1046/j.1523-1747.2000.00023.x

DO - 10.1046/j.1523-1747.2000.00023.x

M3 - Article

C2 - 10886507

AN - SCOPUS:0033939851

VL - 115

SP - 48

EP - 56

JO - Journal of Investigative Dermatology

JF - Journal of Investigative Dermatology

SN - 0022-202X

IS - 1

ER -