Structure, biological activity of the upstream regulatory sequence, and conserved domains of a middle molecular mass neurofilament gene of Xenopus laevis

During development, the molecular compositions of neurofilaments (NFs) unde rgo progressive modifications that correlate with successive stages of axon al outgrowth. Because NFs are the most abundant component of the axonal cyt oskeleton, understanding how these modifications are regulated is essential for knowing how axons control their structural properties during growth. I n vertebrates ranging from lamprey to mammal, orthologs of the middle molec ular mass NF protein (NF-M) share similar patterns of expression during axo nal outgrowth, which suggests that these NF-M genes may share conserved reg ulatory elements. These elements might be identified by comparing the seque nces and activities of regulatory domains among the vertebrate NF-M genes. The frog, Xenopus laevis, is a good choice for such studies, because its ea rly neural development can be observed readily and because transgenic embry os can be made easily. To begin such studies, we isolated genomic clones of Xenopus NF-M(2), tested the activity of its upstream regulatory sequence ( URS) in transgenic embryos, and then compared sequences of regulatory regio ns among vertebrate NF-M genes to search for conserved elements. Studies wi th reporter genes in transgenic embryos found that the 1.5 kb URS lacked th e elements sufficient for neuron-specific gene expression but identified co nserved regions with basal regulatory activity. These studies further demon strated that the NF-M 1.5 kb URS was highly susceptible to positional effec ts, a property that may be relevant to the highly variant, tissue-specific expression that is seen among members of the intermediate filament gene fam ily. Non-coding regions of vertebrate NF-M genes contained several conserve d elements. The region of highest conservation fell within the 3' untransla ted region, a region that has been shown to regulate expression of another NF gene, NF-L. Transgenic Xenopus may thus prove useful for testing further the activity of conserved elements during axonal development and regenerat ion. (C) 2000 Elsevier Science B.V. All rights reserved.