Induction of testis development in mammals requires the presence of the Y-chromosome gene SRY. This gene must exert its effect by interacting with other genes in the sex-determination pathway. Cloning of a translocation chromosome breakpoint from a sex-reversed patient with campomelic dysplasia, followed by mutation analysis of an adjacent gene, indicates that SOX9, an SRY-related gene, is involved in both bone formation and control of testis development.

In humans, mutations in SOX9 result in a skeletal malformation syndrome, campomelic dysplasia (CD). The present study investigated two major classes of CD mutations: 1) point mutations in the high mobility group (HMG) domain and 2) truncations and frameshifts that alter the C terminus of the protein. We analyzed the effect of one novel mutation and three other point mutations in the HMG domain of SOX9 on the DNA binding and DNA bending properties of the protein. The F12L mutant HMG domain shows negligible DNA binding, the H65Y mutant shows minimal DNA binding, whereas the A19V mutant shows near wild type DNA binding and bends DNA normally. Interestingly, the P70R mutant has altered DNA binding specificity, but also bends DNA normally. The effects of the point mutations were interpreted using a molecular model of the SOX9 HMG domain. We analyzed the effects upon transcription of mutations resembling the truncation and frameshift mutations in CD patients, and found that progressive deletion of the C terminus causes progressive loss of transactivation. Maximal transactivation by SOX9 requires both the C-terminal domain rich in proline, glutamine, and serine and the adjacent domain composed entirely of proline, glutamine, and alanine. Thus, CD arises by mutations that interfere with DNA binding by SOX9 or truncate the C-terminal transactivation domain and thereby impede the ability of SOX9 to activate target genes during organ development.In humans, mutations in SOX9 cause campomelic dysplasia (CD), 1 a skeletal malformation syndrome that is often associated with XY sex reversal (1). Other tissues affected include kidney, heart, and brain, consistent with the expression pattern of Sox9 in developing mouse (2, 3). There are four major classes of mutations causing CD: 1) amino acid substitutions in the HMG domain (Fig. 1A), 2) truncations or frameshifts that alter the C terminus of SOX9 (Fig. 1B), 3) mutations at splice junctions, and 4) chromosomal translocations, of which classes 1 and 2 are investigated here. Most CD patients are heterozygous for wild type and mutant alleles of SOX9. CD appears to result from haploinsufficiency; presumably, a critical dose of SOX9 is required to switch on the appropriate genes during development. The present study reports the identification in a CD patient of a novel amino acid substitution mutation (H65Y) in the HMG domain of SOX9. We report the effects of this and three other point mutations (F12L, A19V, and P70R) on the DNA binding and DNA bending activities of the HMG domain.SOX proteins represent a large class of transcription factors related to SRY, the testis-determining factor, through their HMG domains that bind and bend DNA in a sequence-specific manner. Expression of these proteins in defined cell types at specific stages of development appears to govern cell fate decisions. SOX9 activates expression of type II and type XI collagen in vivo (4 -6), consistent with a role in bone development.SOX proteins fall within a larger group of HMG domain proteins comprising two clas...

Cross‐hybridization of the grey seal myoglobin gene to human DNA detected a single human myoglobin gene plus an extensive family of sequences apparently related to the central exon of this gene. The functional human gene is 10.4 kb long and has a haemoglobin‐like three exon/two intron structure with long non‐coding regions similar to its seal homologue. At least 300 bp of 5′‐flanking region are closely homologous between the two genes, with the exception of a divergent purine‐rich region 68‐114 bp upstream of the cap site. A diverged tandem repetitive sequence based on (GGAT)165 is located 1100‐1750 bp upstream from the gene; internal homology units within this sequence suggest sequence homogenization by gene microconversions. A second 33‐bp tandem repeat element in the first intron is flanked by a 9‐bp direct repeat, shares homology with other tandem repetitive elements in the human genome and may represent a novel form of transposable element.

U RNAs are highly abundant small nuclear RNAs involved in the processing of messenger RNA. Most U RNA genes are thought to be transcribed by RNA polymerase II (pol II). However, evidence has recently been presented that U6 RNA genes are transcribed by RNA polymerase III (pol III). In the light of these results it was surprising to find that the 5' flanking region of a mouse U6 RNA gene includes a perfect copy of the octamer sequence motif, ATTTGCAT, found in many RNA polymerase II transcription enhancer elements. In the present study we show that deletion of mouse U6 gene sequences upstream of nucleotide position -217, including the octanucleotide motif, reduces U6 transcription by 90% when assayed in Xenopus laevis oocytes, suggesting the presence of a distant control element. DNase I footprinting of the 5' flanking region of the U6 gene shows protection of the octanucleotide sequence. Moreover, the 5' flanking sequence from -217 to -315 can replace the enhancer of a human U2 RNA gene. We therefore conclude that although U6 RNA genes appear to be transcribed by pol III, they are preceeded by an enhancer-like element which can functionally substitute for the enhancer of a pol II-transcribed U RNA gene.

The pseudoautosomal boundaries are the interface between pseudoautosomal and sex chromosome-specific DNA sequences. We have isolated a gene, PBDX, from the human pseudoautosomal boundary region of Xp. The three exons at the 5' end of PBDX are situated in the pseudoautosomal region immediately downstream of MIC2, whereas the other seven exons are in the X-specific region. Hence, PBDX is inherited in two modes: its 5' end is pseudoautosomally inherited and its 3' end is X-linked. The predicted amino acid sequence of the 540 bp coding region is 48% homologous to 12E7, the product of MIC2. By virtue of its position, PBDX becomes an excellent candidate for the XG blood group gene.