Brian Harding

Studies of the Murine Homolog of the Multiple Endocrine Neoplasia Type 1 (MEN1) Gene, men1

The murine homolog of the multiple endocrine neoplasia type 1 (MEN1) gene (men1), which in humans is associated with tumors of the parathyroids, pancreas, and pituitary, has been characterized by isolating 27 clones from a mouse embryonic stem cell cDNA library. The insert sizes ranged from 600–2500 bp, and sequence analysis identified a 1833 bp open reading frame encoding a 611 amino acid protein. In addition, two clones contained an unspliced intron 1, and another two clones contained 20–29 bp of an upstream sequence, which suggested the presence of an alternate exon 1. This was supported by an analysis of the homologous human sequence. The mouse and human coding regions had 89% and 96% identity of the nucleotide and amino acid sequences, respectively. Investigation of clones isolated from a 129ola mouse genomic library, revealed the men1 gene to consist of 10 exons that spanned ∼6 kb. Northern blot analysis demonstrated the ubiquitous expression of 2.9 kb and 3.4 kb transcripts in mouse adult tissues and embryos from 7 days. DNA sequence analysis of the larger 3.4 kb transcript revealed it to result from a retention of intron 1. In situ hybridization confirmed an early ubiquitous expression in whole mount mouse embryos and adult tissues, but in the latter, different levels of cellular expression were observed, e.g., men1 expression was higher in testicular Sertoli cells than in germ cells. Thus, the mouse men1 gene and the basis of alternative transcripts have been defined, and these will help to facilitate studies of a mouse model.

Mapping the Gene Causing Hereditary Primary Hyperparathyroidism in a Portuguese Kindred to Chromosome 1q22-q31

A Portuguese kindred with autosomal dominant isolated primary hyperparathyroidism (HPT) that was associated with parathyroid adenomas and carcinomas was investigated with the aim of determining the chromosomal location of this gene, designated HPTPort. Leukocyte DNA from 9 affected and 16 unaffected members and 7 parathyroid tumors from 4 patients was used in comparative genomic hybridization (CGH), tumor loss of heterozygosity (LOH), and family linkage studies. The CGH studies revealed abnormalities of chromosomes 1 and 13, and the results of LOH studies were consistent with the involvements of tumor suppressor genes from these regions. Family segregation studies mapped HPTPort to chromosome 1q22‐q31 by establishing linkage with eight loci (D1S254, D1S222, D1S202, D1S238, D1S428, D1S2877, D1S422, and D1S412) (peak two‐point LOD scores = 3. 46–5.14 at 0% recombination), and defined the location of HPT Port to a 21 cM region flanked centromerically by D1S215 and telomerically by D1S306. Thus, HPTPort has been mapped to chromosome 1q22‐q31, and a characterization of this gene will help to elucidate further the mechanisms that are involved in the development of parathyroid tumors.

EagI and NotI linking clones from human chromosomes 11 and Xp

EagI andNotI linking libraries were prepared in the lambda vector, EMBL5, from the mouse-human somatic cell hybrid 1W1LA4.9, which contains human chromosomes 11 and Xp as the only human component. Individual clones containing human DNA were isolated by their ability to hybridise with total human DNA and digested withSalI andEcoRI to identify the human insert size and single-copy fragments. The mean (± SD) insert sizes of theEagI andNotI clones were 18.3 ± 3.2 kb and 16.6 ± 3.6 kb, respectively. Regional localisation of 66 clones (52EagI, 14NotI) was achieved using a panel of 20 somatic cell hybrids that contained different overlapping deletions of chromosomes 11 or Xp. Thirty-nine clones (36EagI, 3NotI) were localised to chromosome 11; 17 of these were clustered in llg13 and another nine were clustered in 11ql4–q23.1. Twenty-seven clones (16EagI, 11NotI) were localised to Xp and 10 of these were clustered in Xpll. The 66 clones were assessed for seven different microsatellite repetitive sequences; restriction fragment length polymorphisms for five clones from 11q13 were also identified. TheseEagI andNotI clones, which supplement those previously mapped to chromosome 11 and Xp, should facilitate the generation of more detailed maps and the identification of genes that are associated with CpG-rich islands.