MOLECULAR PATHOPHYSIOLOGY OF GROWTH DISORDERS

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We have undertaken a patient-based study of the role of molecular defects in individuals with growth disorders. We hypothesize that a subset of individuals with familial severe isolated Growth Hormone (GH) deficiency have a mutation in a gene compromising the Growth Hormone axis. Careful phenotyping via a detailed endocrine evaluation, we limit the potential candidate genes to the Growth Hormone Releasing Hormone (GHRH), the GHRH Receptor (GHRHR), the Growth Hormone Secretagogue (GHS), the GHS receptor (GHSR) and Growth Hormone (GH1). We recruit and analyze families for genetic linkage of familial, severe short stature to a region of the genome containing one of the components of the GH axis. The candidate gene is then characterized, exon- by-exon, initially by Single Strand Conformational Analysis (SSCA) and then by direct sequencing of the conformationally unique exon in affected and unaffected members of a family. Confirmation of mutational status is furnished by restriction fragment length (RFLP) analysis which detects sequence variants, including novel restriction sites. This protocol requires knowledge of the genetic position of a candidate gene, and its fine genetic structure. Those genes whose genomic structure is not known (e.g. GHRHR and GHSR) will be fully characterized by us in an effort to enable their screening as above. We are also recruiting individuals with GH-secreting tumors to determine the role of molecular defects in GH excess/acromegaly. We are revere transcribing the total RNA from tumor tissue to obtain cDNA for candidate genes, including the GHRHR, the GHSR and the Growth Hormone Factor-1 (GHF-1, also called the PIT-1 transcription factor). In cases where RNA is not available, we rely on SSCA followed by direct sequencing of genomic DNA to identify activating mutations resulting in GH excess/acromegaly.