Differential localization of collagen type IX isoform messenger RNAs during early ocular development.

PURPOSE To determine the temporal and spatial localization of the messenger RNAs (mRNAs) for the two collagen alpha 1(IX) isoforms during early development of the embryonic chicken eye. METHODS The reverse transcription-polymerase chain reaction method was used to amplify mRNAs for the two collagen alpha 1(IX) isoforms. mRNA was extracted from optic vesicles of chicken embryos at stages 14 to 19 and from microdissected ocular tissues of older embryonic eyes. After synthesis of complementary DNA, the polymerase chain reaction was performed for 20, 25, or 30 cycles. This ensured a reliable estimate of the relative abundance of the two mRNAs at different stages and in different ocular tissues. Data from the polymerase chain reaction were confirmed by Northern blot analysis. RESULTS mRNA for the shorter alpha 1(IX) chain was present in the optic vesicle as early as stage 14, whereas mRNA for the longer alpha 1(IX) chain was not detectable until after the optic vesicle and lens started to invaginate (stage 15; day 2.5). In later stages of development, mRNAs for both alpha 1(IX) chains were present predominantly in the presumptive ciliary epithelium. They were just detectable in the neural retina at stages 20 and 23 (days 3 to 3.5). By E6 no mRNA for the shorter alpha 1(IX) isoform was detected in the retina, although a trace of the longer alpha 1(IX) isoform was still present. In the lens, mRNA for neither isoform was detectable at any stage. CONCLUSIONS The two isoforms of alpha 1(IX) collagen mRNA are expressed differentially in space and time during early development of the embryonic chicken eye. These molecules may serve as markers for the early specialization of ciliary epithelium as a distinct region of the optic cup.