Spitz François, Benbacer Laïla, Sabourin Jean Christophe, Salminen Marjo, Chen Fengmei, Cywiner Charlotte, Kahn Axel, Chatelet François, Maire Pascal, Daegelen Dominique
INSERM U567, CNRS UMR 8104, Institut Cochin; Department Génétique, Développement et Pathologie Moléculaire, Universiteé René Descartes Paris V, 24 rue du Faubourg Saint Jacques, 75014 Paris.
Differentiation. 2002 Oct;70(8):457-67. doi: 10.1046/j.1432-0436.2002.700808.x.
We have previously shown that the proximal sequences of the human aldolase A fast-muscle-specific promoter (pM) are sufficient to target the expression of a linked CAT reporter gene to all fast, glycolytic trunk and limb muscles of transgenic mice (pM310CAT lines) in a manner mimicking the activity of the endogenous mouse promoter. When a NF1-binding site (motif M2) in this proximal regulatory region is mutated, the activity of the corresponding mM2 transgene is strongly affected but only in a some fast muscles. Here we show that the mutation of the M2 motif has only mild effects on pM activity in axial and proximal limb, while it drastically reduces this activity in both fore and hind limb distal muscles. At the cellular level, we show that both the pM310CAT and mM2 transgenes are highly expressed in fast glycolytic 2B fibers. However, by contrast to the pM310CAT transgene, whose expression is mainly restricted to fast glycolytic 2B fibers, the mM2 transgene is also active in a high proportion of 2X fibers. This result suggests that the M2 sequence could play a role in restricting the expression of pM to the 2B fibers. The variable expression of the mM2 transgene along the limb axis already exists at post-natal day 10 and seems to result from a change in the proportion of expressing fast fibers per muscle. Altogether, these results suggest that, although considered as phenotypically similar, different populations of fast glycolytic fibers exist, in which the requirement of the NF1 activity for pM expression varies according to the proximal versus distal position of the muscle along the limb axis.
我们先前已经表明，人醛缩酶A快肌特异性启动子（pM）的近端序列足以将连接的CAT报告基因的表达靶向转基因小鼠的所有快肌、糖酵解型躯干和四肢肌肉（pM310CAT系），其方式类似于内源性小鼠启动子的活性。当该近端调节区域中的NF1结合位点（基序M2）发生突变时，相应的mM2转基因的活性受到强烈影响，但仅在一些快肌中如此。在此我们表明，M2基序的突变对轴肌和近端肢体中的pM活性仅有轻微影响，而在前后肢远端肌肉中则大幅降低该活性。在细胞水平上，我们表明pM310CAT和mM2转基因在快糖酵解型2B纤维中均高度表达。然而，与主要局限于快糖酵解型2B纤维表达的pM310CAT转基因不同，mM2转基因在高比例的2X纤维中也具有活性。这一结果表明，M2序列可能在将pM的表达限制于2B纤维中发挥作用。mM2转基因沿肢体轴的可变表达在出生后第10天就已存在，似乎是由于每块肌肉中表达快纤维的比例发生了变化。总之，这些结果表明，尽管被认为在表型上相似，但存在不同群体的快糖酵解纤维，其中NF1活性对pM表达的需求根据肌肉沿肢体轴的近端与远端位置而有所不同。
