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J.S. Dalal, A.C. Dose, B. Burnside, B.–A. Battelle; Developmental Expression of Myo3B in Mouse Retina . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3964.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Our long–term goal is to understand the biological functions of class III myosins in the retina. These myosins are unique in having a kinase domain at their N–terminals. Here we report the cloning of two isoforms of myosin3B (myo3B) from adult mouse retina and the postnatal developmental expression of myo3B in mouse retina. Methods: An homology based cloning strategy was used to clone two isoforms of myo3B from adult mouse retina. For analysis of the developmental expression of myo3B, retinas were dissected from animals of different ages at 2 PM in the light. Beginning at p21, animals were dark–adapted for 2 hrs before dissection. In one experiment, animals were maintained in the dark for about 20 hrs and dissected in the dark under dim red light. Myo3b mRNA levels in the retinas of these animals were compared to that in retinas of littermates maintained in cyclic light and dissected in the light. Total RNA from these animals was assayed for quantification of myo3B and opsin mRNA, and 18s rRNA levels by real time PCR using the SYBR green technology (ABI PRISM 7000 sequence detection system). Myo3B was localized in methanol/formaldehyde fixed adult retinas using a polyclonal antibody raised in rabbits that was directed against a 209 amino acid long C–terminal fragment of Myo3B and expressed in E. coli as a fusion protein. Results: Two Myo3B isoforms are expressed in the mouse retina. Sequence analysis showed that the mouse Myo3B is 92% identical to human Myo3B in the kinase domain, 91% in motor domain, and 39% in the tail domain. Real time PCR analysis of various adult tissues showed that myo3B is highly expressed in retina, testis and olfactory bulb. Lower but detectable levels were found in brain, kidney and thymus. During postnatal retinal development, myo3B mRNA levels increased during the first three weeks after birth then decreased to adult levels by about four weeks. Long–term dark adaptation did not influence myo3B mRNA levels in adult retinas. Antibody staining of the retinas showed that Myo3B is concentrated in the inner segments of the photoreceptors of adult animals. Conclusions: Two isoforms of Myo3B are expressed in mouse retina. Myo3B is also expressed in mouse kidney, testis, brain, olfactory bulb and thymus. Myo3B expression peaks at p21 (eye opening) in retina. Light does not influence myo3B expression in the adult mouse retina. Preliminary results show that Myo3B is expressed in the inner segments of mouse photoreceptors.
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