Survival Motor Neuron (SMN) polymorphism in relation to congenital arthrogryposis in Piedmont cattle

In farm animals candidate gene approach can be a useful tool to identify disease genes which afterwards can be included in selection programme analysis. The term arthrogryposis refers to a symptom complex that is characterised by congenital contractures. It consists of different degrees of flexion rigidity of phalangeal, metacarpophalangeal, carpal and metatarsophalangeal joints, Arthrogryposis has been reported in man, in farm animals and in pets. Several forms have been reported having a genetic origin in man. Recently in a pedigree study it has been indicated an autosomal recessive condition with congenital arthrogryposis in Suffolk lambs. In Brown Swiss and Holstein Friesian cattle congenital contractures associated with degeneration and loss of motor neurons in spinal cord, have been recorded and classified as Spinal Muscular Atrophy (SMA). Survival Motor Neuron gene (SMN) has been suggested as candidate gene for SMA. In man SMN shows a centromeric and a telomeric copy. Several exon or gene duplication deletions or point mutations, particularly in exon 7 and 8, have been recorded associated to arthrogryposis. In German Brown SMN cDNA has been sequenced (Genbank#AF035323) and mapped by Pietrowsky et al. in 1999 on chr20q12-13, by BLAST it shows high identity with human sequence. Eggen et al. in 1999 defined SMN mapping on chr20q14. Linkage between SMN and SMA has not yet identified in cattle. In the last 20 years National Association of Piedmont Cattle recorded a mean of 1.8% arthrogryposis cases. Incidence of 0.6% in females and 3.3% in males suggests the hypothesis of genetic disorder with incomplete penetrance in male. In our study we cloned and sequenced SMN gene starting from cDNA extracted from spinal cord samples of 2 animals: one Piedmont calf showing a severe clinical form of arthrogryposis and a normal Piedmont calf as a control. RT-PCR has been performed by a couple of primers we designed to amplify exon 1-6 region. Other primers have been designed to amplify, coupled with commercial adapter primers, the 5'end in a 5'RACE (Rapid Amplification of cDNA Ends). PCR products have been cloned in E. Coli by plasmid vector, sequenced by Big Dye Terminators chemical (Applied Biosystems) and analysed by BLAST® and Navigator software (Applied Biosysterns). By comparing the sequence available in Genbank with the sequences we obtained by RT-PCR, a silent substitution C->A in exon 6, both from health and affected calf, has been shown. By 5'RACE, > 50% of the clones in the affected calf showed a Single Nucleotide Polymorphism (SNP) ATG->TTG in exon 1 that should determine an aminoacid change Met->Leu (single point mutation M3L). In our clones this mutation is 100% associated to 9bp increase length of 5'UTR and to a silent mutation TTC-> TTT in exon 1 (Genbank#AF466194). No single point mutation or 5' end polymorphism have been shown in healthy control and in the remaining 50% of clones from the affected calf (Genbank#AF466193). 5'RACE PCR products from the affected animal, which were previously cloned, have been also directly sequenced to control the co-presence in the same product of both the two different sequences. Also in children SMA- affected an A2G point mutation associate to 5'UTR polymorphism has been described. Therefore it is possible to assume a pathogenic effect of exon 1 mutations both in man and cattle, as they are located in the SMN region where SP1 interacts to form the spliceosoma, a fundamental complex for cell splicing mechanism. Studies are in progress to assess this assumption in man (V.Funanage personal communication). Research supported by R.A.I.Z. (N. RZ-271). (Texte intégral)

Bibliographic Details

Main Authors:	Longeri, M., Perrone, Trina, Bongioni, G., Bona, M., Zanotti, M., Galli, A.
Format:	conference_item biblioteca
Language:	eng
Published:	CIRAD
Subjects:	L10 - Génétique et amélioration des animaux, polymorphisme génétique, maladie des articulations, bétail, http://aims.fao.org/aos/agrovoc/c_24031, http://aims.fao.org/aos/agrovoc/c_4051, http://aims.fao.org/aos/agrovoc/c_4397, http://aims.fao.org/aos/agrovoc/c_4026,
Online Access:	http://agritrop.cirad.fr/512107/
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