Assembly properties of dominant and recessive mutations in the small mouse neurofilament (NF-L) subunit.
We have generated a set of amino- and carboxy-terminal deletions of the NF-L neurofilament gene and determined the assembly properties of the encoded subunits after coexpression with vimentin or wild-type NF-L. NF-L molecules missing greater than 30% (31 amino acids of the head) or 90% (128 amino acids of the tail) failed to incorporate into intermediate filament networks. Carboxy-terminal deletions into the rod domain yield dominant mutants that disrupt arrays assembled from wild-type subunits, even when present at levels of approximately 2% of the wild-type subunits. Even mutants retaining 55% of the tail (61 amino acids) disrupt normal arrays when accumulated above approximately 10% of wild-type subunits. Since deletion of greater than 90% of the head domain produces "recessive" assembly incompetent subunits that do not affect wild-type filament arrays, whereas smaller deletions yield efficient network disruption, we conclude that some sequence(s) in the head domain (within residues 31-87) are required for the earliest steps in filament assembly. Insertional mutagenesis in the nonhelical spacer region within the rod domain reveals that as many as eight additional amino acids can be tolerated without disrupting assembly competence.
Gill
S R
SR
Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Wong
P C
PC
Monteiro
M J
MJ
Cleveland
D W
DW
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002872:Chromosome Deletion; D005455:Fluorescent Antibody Technique; D005799:Genes, Dominant; D005808:Genes, Recessive; D007381:Intermediate Filament Proteins / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D007382:Intermediate Filaments / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D007739:L Cells; D051379:Mice; D008969:Molecular Sequence Data; D016900:Neurofilament Proteins; D013329:Structure-Activity Relationship; D014162:Transfection
D007381:Intermediate Filament Proteins; D016900:Neurofilament Proteins