Next-generation sequencing identifies the Danforth's short tail mouse mutation as a retrotransposon insertion...

Area of Science:

• Developmental Biology
• Genetics
• Molecular Biology

Background:

• The semidominant Danforth's short tail (Sd) mutation in mice causes severe axial skeleton malformations, absent tail, kidney agenesis, and persistent cloaca.
• These phenotypes resemble human caudal malformation syndromes, including urorectal septum malformation and VACTERL association.
• Previous mapping localized the Sd mutation to a 0.9 cM region on mouse chromosome 2qA3.

Purpose of the Study:

• To identify the causative genetic mutation responsible for the Danforth's short tail (Sd) phenotype.
• To understand the molecular mechanisms underlying caudal malformations in Sd mutant embryos.
• To provide a genetic model for studying human caudal malformation syndromes.

Main Methods:

• Sanger sequencing of exons and intron/exon boundaries within the Sd critical region.
• DNA enrichment/capture followed by next-generation sequencing (NGS) of the critical genomic region.
• Bioinformatic analysis of NGS data, including interrogation of discarded reads.
• Analysis of Ptf1a gene expression in Sd mutant embryos.

Main Results:

• Sanger sequencing and standard NGS analysis did not reveal causative mutations.
• Analysis of discarded NGS reads identified an early transposon (ETn) retroviral insertion 12.5 kb upstream of the Ptf1a gene.
• Ptf1a gene expression was significantly upregulated in Sd mutant embryos at embryonic day 9.5 (E9.5).

Conclusions:

• An ETn retroviral insertion upstream of the Ptf1a gene is identified as the causative mutation for the Danforth's short tail (Sd) phenotype.
• This insertion leads to upregulation of Ptf1a expression, contributing to the observed developmental defects.
• The Sd mouse model offers a valuable tool for investigating the genetic basis and developmental pathways of human caudal malformation syndromes.