Restriction analysis of otosclerosis-associated CD46 splicing variants

Otosclerosis is a primary bone remodeling disorder of the human otic capsule and is associated with persistent measles virus infection. The human cellular receptor of measles virus is the membrane cofactor protein (MCP, CD46), which has 14 well-described splicing variants. Unique CD46 expression pattern of the otic capsule and the stapes footplate may determine the susceptibility for persistent measles virus infection. A total of 51 surgically removed ankylotic stapes footplates were analyzed by histopathological and molecular biological methods, respectively. Nucleic acids were extracted. Measles virus sequences were detected by nucleoprotein RNA-speciWc reverse transcriptase polymerase chain reaction (RT-PCR). Alternatively spliced RNA of CD46 isoforms was ampli- Wed by RT-PCR; cDNA amplimers were separated by SDS poly-acrylamide gel electrophoresis and were puriWed from the gel. Complementary DNA of CD46 isoforms was restricted by endonuclease enzymes having CD46-speciWc recognition sites. The presence of viral RNA was associated exclusively with the histopathological diagnosis of otosclerosis; the stapes specimens with negative measles virus belonged to non-otosclerotic stapes Wxations. All specimens (N = 51) were characterized by the consecutive expression of Wve CD46 variants (c, d, e, f and one shorter unidentiWed isoform). Histologically conWrmed ostosclerotic specimens (N = 21) were characterized by increased expression levels of variant "f" and the unknown isoform. Increased expression levels of these isoforms and special CD46 expression pattern of the human otic capsule might produce modiWed or pathological intracellular signalization that could create the possibility of persistent measles virus infectio