Definition of the mutation responsible for maple syrup urine disease in poll shorthorns and genotyping poll shorthorns and poll herefords for maple syrup urine disease alleles

The organisation of the E1 alpha(l) subunit of bovine branched-chain alpha- keto acid dehydrogenase gene was established. CDNA was cloned from Poll Sho rthorn x Poll Hereford calves affected with Maple Syrup Urine Disease to id entify the mutation responsible for the disease in Poll Shorthorns. Clones containing the cDNA sequences inherited from the Poll Shorthorn sire of the affected calves were identified. Paternal clones were sequenced and a cyti dine to thymidine transition was found at nucleotide 1380. The mutation is predicted to substitute leucine in place of a highly conserved proline at c odon 372. A polymerase chain reaction procedure was developed for detection of the 1380C-->T mutation in genomic DNA. Three Poll Shorthorn parents of affected calves and three affected Poll Shorthorn x Poll Hereford calves we re heterozygous and an affected Poll Shorthorn calf was homozygous for this mutation. An improved polymerase chain reaction procedure was also devised to genotype Poll Herefords for the 248C-->T mutation. The proce dures will facilitate disease prevention programs and assist in differential diagnosi s of conditions in new-born calves that present with a rapid onset of progr essive neurological disease and are characterised histologically by 'status spongiosus'. Maple Symp Urine Disease (MSUD) is an autosomal recessive def ect reported in humans (Danner and Elsas 1989), and in Poll Hereford (PH) a nd Poll Shorthorn tps) calves (Harper et al 1986, Healy et al 1992). The cl inical, biochemical and pathological manifestations of the disease are iden tical in the two breeds of cattle, and are characterised by the rapid onset of progressive neurological disease, leading to death within a few days of birth. The disease is caused by a deficiency of activity of the mitochondr ial enzyme branched-chain alpha-keto acid dehydrogenase (BCKADH). This defi ciency leads to elevated concentrations, in blood and tissues, of branched chain alpha-keto acids and their precursors, the branched chain amino acids , valine, leucine and isoleucine. BCKADH consists of four subunits E1 alpha , E1 beta, E2 and E3 that are encoded by separate genes, and MSUD may resul t from deficiency of any of the subunits. In PHS, the disease in caused by premature termination of translation, of the E1 alpha subunit, that is indu ced by a cytidine to thymidine transition exon 2 (248C-->T), that converts the glutamine codon -6 to a stop codon (Q-6ST; Zhang et al 1990). We have s hown that MSUD-affected ps x PH calves are heterozygous at the PH locus, il lustrating molecular heterogeneity exists for bovine MSUD (Healy and Dennis 1994a). The fact that these crossbred calves are affected, indicates the p s, like the PH mutation, resides in the E1 alpha subunit.