ISOLATED SEQUENCES FROM THE LINKED MYF-5 AND MRF4 GENES DRIVE DISTINCT PATTERNS OF MUSCLE-SPECIFIC EXPRESSION IN TRANSGENIC MICE

In developing mouse embryos, MyoD family regulatory genes are expresse d specifically in muscle precursors and mature myofibers. This pattern , taken together with the well-established ability of MyoD family memb ers to convert a variety of cell types to skeletal muscle, suggests a significant role for these genes in regulating skeletal myogenesis. Th e possibility that expression of these genes may be causally associate d with segregation of the myogenic lineage from other mesodermal deriv atives, or with the subsequent maintenance of muscle phenotypes at lat er times, raises the issue of how MyoD family genes are themselves reg ulated during development. In this work, we have initiated studies to identify DNA sequences that govern Myf-5 and MRF4 (herculin, myf-6) tr anscription. Myf-5 is the first of the MyoD family to be expressed in the developing mouse embryo, while MRF4 is the most abundantly express ed myogenic factor in postnatal animals. In spite of their strikingly divergent patterns of expression, Myf-5 and MRF4 are tightly linked in the mouse genome; their translational start codons are only 8.5 kilob ases apart. Here, the 5' flanking regions of the mouse Myf-5 and MRF4 genes were separately linked to a bacterial beta-galactosidase (lacZ) gene, and these constructs were each used to produce several lines of transgenic mice. Transgene expression was monitored by X-gal staining of whole embryos and by in situ hybridization of embryo sections. For the Myf-5/lacZ lines, the most intense transgene expression was in the visceral arches and their craniofacial muscle derivatives, beginning at day 8.75 post coitum (p.c.). This correlates with endogenous Myf-5 expression in visceral arches. However, while Myf-5 is also expressed in somites starting at day 8 p.c., transgene expression in the trunk i s not observed until day 12 p.c. Thus, the Myf-5/lacZ construct respon ds to early Myf-5 activators in the visceral arches but not in the som ites, suggesting that myogenic determination in the nonsomitic head me soderm may be under separate control from that of the somitic trunk me soderm. MRF4/lacZ lines displayed an entirely different pattern from M yf-5. Transgene expression appeared in muscles starting at day 16.5 p. c. and became increasingly prominent at later times. However, an early wave of myotomal expression that is characteristic of the endogenous MRF4 was not recapitulated by the transgene.