TY - JOUR
T1 - Expression of tyrosinase and the tyrosinase related proteins in the Mitf(vit) (Vitiligo) mouse eye
T2 - Implications for the function of the microphthalmia transcription factor (Mitf)
AU - Smith, Sylvia B.
AU - Zhou, B. K.
AU - Orlow, Seth J.
N1 - Funding Information:
Supported by NIH (EY09682 to SBS and EY10223 to SJO), by the Medical College of Georgia Research Institute and by an unrestricted grant from Research to Prevent Blindness, Inc. New York, to the Department of Ophthalmology, Medical College of Georgia. The C57BL}6-Mitfvit mice were the offspring from our colony of breeding pairs that were provided by Dr R. L. Sidman, New Eng. Regional Primate Res. Ctr., Southboro, MA, U.S.A. (funded by NEI grant EY06859). We thank Dr Vincent J. Hearing for the gift of the antibodies against tyrosinase, TRP-1 and TRP-2, Sharon Pifko-Hirst for expert technical assistance and Doris McCool for care and maintenance of the mouse colony and Ms Penny Roon for assistance with the immunohistochemical studies.
PY - 1998/4
Y1 - 1998/4
N2 - Mitf (Microphthalmia transcription factor), a basic-helix-loop-helix zipper protein, encoded at the microphthalmia (Mitf) locus, regulates the transcription of the gene encoding tyrosinase, the rate-limiting enzyme in melanin biosynthesis, by binding the DNA sequence CATGTG. This binding site is present also in the genes encoding two tyrosinase related proteins, TRP-1 and TRP-2. To gain insight into the function of Mitf in vivo, we determined whether there was a difference in the levels of these proteins in the RPE/choroid of the vitiligo (Mitf(vit)) mouse, in which there is a mutation of the Mitf gene. This mouse has alteration of RPE pigmentation and function that presumably leads to slow progressive loss of photoreceptor cells. The RPE/choroid was dissected from eyes of vitiligo and C57BL/6 wild-type mice at postnatal ages 2, 4, 7, 10, 14, 21 and 42 days. Extracts of pooled tissues were subjected to electrophoresis and immunoblotting. The levels of tyrosinase, TRP-1 and TRP-2 were determined densitometrically following immunodetection with rabbit antipeptide antisera. In addition, the tyrosine hydroxylase activity of tyrosinase as assayed radiometrically. Levels of TRP- 1 were 3-7 fold greater in control RPE/choroid compared with mutants. This marked difference in protein level was observed at the earliest age examined (P2) and persisted throughout the first two weeks. Tyrosinase levels in mutants were similar to controls at P2 and P4, but were reduced at P10 and beyond. Tyrosinase activity was diminished also in mutants by P10. Levels of TRP-2 were similar between mutants and controls, although the typical decrease seen in controls after P14 was attenuated in the mutant mice. There is a significant reduction in the level of TRP-1 in the RPE/choroid of the Mitf(vit) mouse. The data suggests that transcription of the gene encoding TRP-1 is extremely dependent upon functional Mitf. It provides in vivo evidence that Mitf regulates the transcription of the gene encoding TRP-1 as well as tyrosinase.
AB - Mitf (Microphthalmia transcription factor), a basic-helix-loop-helix zipper protein, encoded at the microphthalmia (Mitf) locus, regulates the transcription of the gene encoding tyrosinase, the rate-limiting enzyme in melanin biosynthesis, by binding the DNA sequence CATGTG. This binding site is present also in the genes encoding two tyrosinase related proteins, TRP-1 and TRP-2. To gain insight into the function of Mitf in vivo, we determined whether there was a difference in the levels of these proteins in the RPE/choroid of the vitiligo (Mitf(vit)) mouse, in which there is a mutation of the Mitf gene. This mouse has alteration of RPE pigmentation and function that presumably leads to slow progressive loss of photoreceptor cells. The RPE/choroid was dissected from eyes of vitiligo and C57BL/6 wild-type mice at postnatal ages 2, 4, 7, 10, 14, 21 and 42 days. Extracts of pooled tissues were subjected to electrophoresis and immunoblotting. The levels of tyrosinase, TRP-1 and TRP-2 were determined densitometrically following immunodetection with rabbit antipeptide antisera. In addition, the tyrosine hydroxylase activity of tyrosinase as assayed radiometrically. Levels of TRP- 1 were 3-7 fold greater in control RPE/choroid compared with mutants. This marked difference in protein level was observed at the earliest age examined (P2) and persisted throughout the first two weeks. Tyrosinase levels in mutants were similar to controls at P2 and P4, but were reduced at P10 and beyond. Tyrosinase activity was diminished also in mutants by P10. Levels of TRP-2 were similar between mutants and controls, although the typical decrease seen in controls after P14 was attenuated in the mutant mice. There is a significant reduction in the level of TRP-1 in the RPE/choroid of the Mitf(vit) mouse. The data suggests that transcription of the gene encoding TRP-1 is extremely dependent upon functional Mitf. It provides in vivo evidence that Mitf regulates the transcription of the gene encoding TRP-1 as well as tyrosinase.
KW - Microphthalmia transcription factor
KW - Mitf
KW - Pigmentation genes
KW - Retinal degeneration
KW - Retinal pigment epithelium
KW - TRP-1
KW - Tyrosinase
KW - Vitiligo mouse
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U2 - 10.1006/exer.1997.0443
DO - 10.1006/exer.1997.0443
M3 - Article
C2 - 9593634
AN - SCOPUS:0032055073
SN - 0014-4835
VL - 66
SP - 403
EP - 410
JO - Experimental eye research
JF - Experimental eye research
IS - 4
ER -