Defining the Properties of the Nonhelical Tail Domain in Type II Keratin 5: Insight from a Bullous Disease-causing Mutation

Li-Hong Gu and Pierre A. Coulombe*

Departments of Biological Chemistry and Dermatology, The Johns Hopkins University School of Medicine, Baltimore, MD 21205

Monitoring Editor: Thomas Pollard

Inherited mutations in theintermediate filament (IF) proteins keratin 5 (K5) or keratin14 (K14) cause epidermolysis bullosa simplex (EBS), in whichbasal layer keratinocytes rupture upon trauma to the epidermis.Most mutations are missense alleles affecting amino acids locatedin the central ${alpha}$ -helical rod domain of K5 and K14. Here we studythe properties of an unusual EBS-causing mutation in which anucleotide deletion (1649delG) alters the last 41 amino acidsand adds 35 residues to the C-terminus of K5. Relative to wildtype,filaments coassembled in vitro from purified K5-1649delG andK14 proteins are shorter and exhibit weak viscoelastic propertieswhen placed under strain. Loss of the C-terminal 41 residuescontributes to these alterations. When transfected in culturedepithelial cells, K5-1649delG incorporates into preexistingkeratin IFs and also forms multiple small aggregates that oftencolocalize with hsp70 in the cytoplasm. Aggregation is purelya function of the K5-1649delG tail domain; in contrast, thecloned 109 residue-long tail domain from wildtype K5 is distributedthroughout the cytoplasm and colocalizes partly with keratinIFs. These data provide a mechanistic basis for the cell fragilityseen in individuals bearing the K5-1649delG allele, and pointto the role of the C-terminal 41 residues in determining K5’sassembly properties.

^*Corresponding author. E-mail: coulombe{at}jhmi.edu