Context. The LHX4 LIM-homeodomain transcription factor has essential roles in pituitary gland and nervous system development. Heterozygous mutations in LHX4 are associated with combined pituitary hormone deficiency (CPHD).

Objective. To determine the nature and frequency LHX4 mutations in patients with pituitary hormone deficiency and to examine the functional outcomes observed mutations.

Design. The LHX4 gene sequence was determined from patient DNA. The biochemical and gene regulatory properties of aberrant LHX4 proteins were characterized using structural predictions, pituitary gene transcription assays, and DNA binding experiments.

Patients. 253 patients from 245 pedigrees with GH deficiency and typically deficiency of at least one additional pituitary hormone.

Results. In five patients, three types of heterozygous missense mutations in LHX4 that result in substitution conserved amino acids were identified. One substitution is between the LIM domains (R84C); the others are in the homeodomain (L190R; A210P). The patients have GH deficiency; some also display reductions in TSH, LH, FSH, or ACTH and aberrant pituitary morphology. Structural models predict that the aberrant L190R and A210P LHX4 proteins would have impaired DNA binding and gene activation properties. Consistent with these models, electrophoretic mobility shift assays and transfection experiments using pituitary gene promoters demonstrate that whereas the R84C form has reduced activity, the L190R and A210P proteins are inactive.

Conclusions. LHX4 mutations are a relatively rare cause CPHD. This report extends the range phenotypes associated with LHX4 gene mutations and describes three novel exonic mutations in the gene.