Characterization of the Genetic Basis for Autosomal Recessive Hereditary Nephropathy in theEnglishSpringerSpaniel

Abstract

Background: Autosomal recessive hereditary nephropathy (ARHN) was diagnosed in 2 English Springer Spaniels (ESS), a breed not previously reported to be affected by hereditary nephropathy (HN).Objective: To identify and characterize the genetic cause of ARHN in ESS. Animals: Sixty-three ESS (2 with ARHN, 2 obligate carriers, and 59 others), 2 mixed-breed dogs with X-linked HN, and 2 English Cocker Spaniels (ECS) with ARHN were included.Methods: ARHN was diagnosed based on transmission electron microscopy and immunostaining of kidney. DNA from affected dogs was screened for the mutation known to cause ARHN in ECS. Quantities of COL4A3, COL4A4, and COL4A5 mRNA transcripts in renal cortex were determined using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) for ARHN-affected dogs and 7 other dogs. The coding regions of COL4A3 and COL4A4 were sequenced for the 2 ARHN-affected ESS and an unaffected dog. Exon 30 of COL4A4 was sequenced for all 63 ESS.Results: qRT-PCR indicated a significant reduction in transcript levels of both COL4A3 and COL4A4 mRNA in the kidney of ARHN-affected ESS. Sequencing identified a single nucleotide substitution in COL4A4 at base 2806 resulting in a premature stop codon. Thirteen of 25 related dogs were identified as carriers.Conclusions and Clinical Importance: A mutation highly likely to cause ARHN in ESS has been identified.

Kidney diseases caused by glomerular basement membrane typeIVcollagen defects in dogs

“…To date, 4 different examples of canine glomerular diseases caused by identified genetic type IV collagen defects have been recognized (Table ). Two are X‐linked and are caused by different COL4A5 mutations, and 2 are autosomal recessive and are caused by different COL4A4 mutations . However, all 4 mutations cause disruptions of gene translation leading to an inability to synthesize either complete α4(IV) or complete α5(IV) chains and thus preventing the assembly of the α3.α4.α5(IV) network that is crucial for maintenance of GBM structure and function in the adult kidney.…”

“…Dogs with the autosomal recessive forms of HN that have been identified to date are homozygous for COL4A4 mutations that preclude synthesis of complete α4‐chains, so they also cannot form intracellular α3.α4.α5 heterotrimers or assemble a α3.α4.α5 network in their GBM. Although the onset and evolution of ultrastructural changes in their GBM has not been studied as thoroughly in these dogs as it has been in male dogs with XL‐HN, the events that are initiated by the absence of the α3.α4.α5 network are presumed to be similar in both genetic forms of the disease.…”

“…Indeed, the general pattern of relatively mild changes at early stages of the disease that progress to more severe changes as the disease advances was demonstrated in English Cocker Spaniels in which serial renal biopsies were evaluated at 6 and 13 months of age in addition to when the dogs were euthanized due to the development of renal failure . Of note, α5.α5.α6 heterotrimers can be synthesized in dogs with AR‐HN and therefore α5.α5.α6 networks are formed in these dogs not only in the basement membranes where they are universally present in normal dogs (eg, Bowman's capsule and epidermal basement membranes) but also in their GBM . Thus, although dogs with AR‐HN can substitute both the α1.α1.α2 network and the α5.α5.α6 network for the missing α3.α4.α5 network in their GBM, they develop a nephropathy that is clinically and pathologically similar (except for the differences in collagen IV α‐chain content of the various renal basement membranes that can be demonstrated by immunostaining) from the nephropathy that develops in male dogs with XL‐HN that can substitute only the α1.α1.α2 network for the missing α3.α4.α5 network in their GBM.…”

“…Affected dogs appear healthy as puppies, and develop persistent proteinuria of glomerular origin as their first clinical manifestation of disease . The onset of proteinuria most often is at 4–6 months of age, but can be as early as 2–3 months or as late as 6–8 months of age.…”

Emerging perspectives on hereditary glomerulopathies in canines

“…The mutation found was a non-synonymous SNP in exon 30 of COL4A4, changing C to T (c.2712C.T), causing a premature stop codon. 36 A specific PCR test is available (http://www.pawprintgenetics.com).…”