Small cell lung cancer (SCLC) is known to express the
HuD protein, the neuronal
antigen homologous to Drosophila Elav and Sxl genes involved in neuronal and sex development. HuD is the target of an immune response including high titered
antibodies causing
paraneoplastic encephalomyelitis and sensory neuropathy. Because the p53 recessive oncogene is mutated and anti-p53
antibodies frequently occur in
cancer patients, we wondered if the development of anti-HuD
antibodies signaled the presence of HuD mutations in
lung cancer. The HuD gene was mapped to chromosome region 1p using a human-mouse hybrid cell panel. We confirmed that 26 of 46
cancer (43
lung cancer and 3
mesothelioma) cell lines expressed HuD
mRNA and that this expression, as well as
protein expression by Western blot, correlated strongly with the SCLC neuroendocrine phenotype. Southern blot and single-strand conformation polymorphism analyses showed that HuD was not mutated in 78
lung cancers, including patients with the severe
paraneoplastic syndrome. Northern blot analysis showed that
lung cancer cell lines expressed two major mRNAs (4.3 and 4.0 kilobases) of HuD. We found the three previously described alternative spliced
mRNA forms (HuDpro, HuD, and HuDmex). In addition, we also found HuD
mRNA had an alternative splicing form in its 5'-coding region. This alternative splice introduced 87 base pairs of sequence and a
termination codon resulting in a predicted small, truncated
protein (11
amino acids) reminiscent of the male-specific truncated
protein in the Sex-lethal (Sxl) gene of Drosophila. However, mRNAs encoding both full-length and truncated
proteins were expressed in all SCLCs. These results show that the HuD gene is not mutated in
lung cancer, including
tumors from patients producing anti-HuD
antibodies, but HuD expression is an independent marker or determinant of the neuroendocrine differentiation seen in SCLC.