An inversion involving the mouse Shh locus results in brachydactyly through dysregulation of Shh expression

Abstract

Short digits (Dsh) is a radiation-induced mouse mutant. Homozygous mice are characterized by multiple defects strongly resembling those resulting from Sonic hedgehog (Shh) inactivation. Heterozygous mice show a limb reduction phenotype with fusion and shortening of the proximal and middle phalanges in all digits, similar to human brachydactyly type A1, a condition caused by mutations in Indian hedgehog (IHH). We mapped Dsh to chromosome 5 in a region containing Shh and were able to demonstrate an inversion comprising 11.7 Mb. The distal breakpoint is 13.298 kb upstream of Shh, separating the coding sequence from several putative regulatory elements identified by interspecies comparison. The inversion results in almost complete downregulation of Shh expression during E9.5-E12.5, explaining the homozygous phenotype. At E13.5 and E14.5, however, Shh is upregulated in the phalangeal anlagen of Dsh/+ mice, at a time point and in a region where WT Shh is never expressed. The dysregulation of Shh expression causes the local upregulation of hedgehog target genes such as Gli1-3, patched, and Pthlh, as well as the downregulation of Ihh and Gdf5. This results in shortening of the digits through an arrest of chondrocyte differentiation and the disruption of joint development.

Forced chondrocyte expression of sonic hedgehog impairs joint formation affecting proliferation and apoptosis

“…The fused phalangy phenotype was also shown in radiationinduced mouse mutant, called short digit (Dsh), with an inversion on chromosome 5 that resulted in a biphasic regulation of Shh expression (Niedermaier et al, 2005). These mice, in addition to fused phalangy, also displayed short digits, a phenotype that resembles human brachydactyly type A1.…”

“…During rat joint formation, although cavitation occurred within the interzone, no apoptosis was observed, but a rearrangement of this area with incorporation into the outer layers of the central cells (Ito and Kida, 2000). Nonetheless, apoptosis was evidenced during murine interphalangeal joint formation at E14.5 (Niedermaier et al, 2005).…”

“…These mice, in addition to fused phalangy, also displayed short digits, a phenotype that resembles human brachydactyly type A1. In Col2-SHH however, phalangy are not shorter than WT, most likely because the overexpression of Shh, under the control of type II collagen promoter has already occurred at E9.5 (Zhou et al, 1995), whereas in the Dsh mice Shh overexpression was observed at later stages, E13.5-E14.5 (Niedermaier et al, 2005).…”

“…This region is a specialized area of higher cell density that subsequently evolves into a three-layered area consisting of two outer regions of higher cell density and a central region of lower cell density (Archer et al, 2003). Cells, in this central region, first lose properties of chondrogenic cells, then become flattened, stop to produce cartilage extracellular matrix components and eventually die according to some authors (Niedermaier et al, 2005;Spitz and Duboule, 2001) or become incorporated in the outer layers according to others (Wu et al, 2002). These processes lead to the joint cavity formation.…”

Shh and Gremlin1 chromosomal landscapes in development and disease

“…translocations, deletions) affecting this 1 Mb Shh chromosomal landscape should disrupt Shh expression and result in phenotypes such as holoprosencephaly (HPE) and limb truncations, which is indeed the case in mouse embryos homozygous for the Dsh (short digits) mutation. The radiation-induced Dsh mutation is about 11.7 Mb inversion with one of its breakpoints between the Shh and Lmbr1 genes, which not only disrupts the chromosomal landscape but also separates the Shh transcription unit from the cis-regulatory elements [27 ].…”