Perälä M, Savontaus M, Metsäranta M, Vuorio E
Department of Medical Biochemistry and Molecular Biology, University of Turku, Kiinamyllynkatu 10, FIN-20520 Turku, Finland.
Biochem J. 1997 May 15;324 ( Pt 1)(Pt 1):209-16. doi: 10.1042/bj3240209.
Several techniques were used to study the co-ordination of mRNA levels for five constituent chains of cartilage collagen fibrils during mouse development. Short cDNA clones were first constructed for mouse and human alpha3(IX) and for mouse proalpha1(XI) collagen mRNA species. Northern analysis of developing mouse embryos revealed that the mRNA species for alpha1, alpha2 and alpha3 chains of type IX collagen peaked earlier than those for proalpha1(II) and proalpha1(XI) collagen chains. Quantification of these mRNA species by slot-blot hybridization confirmed this developmental regulation: the mRNA ratios for type II/type IX/type XI collagens changed from 5.7:1:0.6 (at embryonic day 12.5) to 10.6:1:0.9 (in newborn mice). However, the genes coding for the three chains of type IX collagen seemed to be under more co-ordinated regulation during mouse development. In addition to high mRNA levels in cartilages and the eye, low levels of type IX collagen transcripts were identified in brain and skin of newborn mouse using RNase protection and reverse transcriptase-PCR assays. Finally, hybridization in situ revealed identical tissue distributions of the three type IX collagen mRNA species during early chondrogenesis but somewhat more widespread expression of the alpha1(IX) and alpha3(IX) mRNA species during endochondral ossification at day 16.5 of embryonic development. These results suggest a relatively tight co-ordination of the alpha1(IX), alpha2(IX), and alpha3(IX) collagen mRNA species in chondrocytes, but a lack of co-ordination in several non-cartilaginous tissues.
运用了多种技术来研究小鼠发育过程中软骨胶原纤维的五条组成链的mRNA水平的协调性。首先构建了小鼠和人类α3(IX)以及小鼠原α1(XI)胶原mRNA种类的短cDNA克隆。对发育中的小鼠胚胎进行Northern分析发现，IX型胶原的α1、α2和α3链的mRNA种类峰值出现得比原α1(II)和原α1(XI)胶原链的mRNA种类更早。通过狭缝印迹杂交对这些mRNA种类进行定量分析证实了这种发育调控：II型/IX型/XI型胶原的mRNA比例从5.7:1:0.6（胚胎第12.5天）变为10.6:1:0.9（新生小鼠）。然而，在小鼠发育过程中，编码IX型胶原三条链的基因似乎受到了更协调的调控。除了在软骨和眼睛中mRNA水平较高外，利用核糖核酸酶保护和逆转录聚合酶链反应分析在新生小鼠的脑和皮肤中也鉴定出了低水平的IX型胶原转录本。最后，原位杂交显示在软骨形成早期，三种IX型胶原mRNA种类具有相同的组织分布，但在胚胎发育第16.5天软骨内骨化过程中，α1(IX)和α3(IX)mRNA种类的表达分布更广。这些结果表明软骨细胞中α1(IX)、α2(IX)和α3(IX)胶原mRNA种类之间存在相对紧密的协调性，但在一些非软骨组织中缺乏协调性。
