Identification of a functional enhancer variant within the chronic pancreatitis‐associatedSPINK1c.101A>G (p.Asn34Ser)‐containing haplotype

Abstract

The haplotype harboring the SPINK1 c.101A>G (p.Asn34Ser) variant (also known as rs17107315:T>C) represents the most important heritable risk factor for idiopathic chronic pancreatitis identified to date. The causal variant contained within this risk haplotype has however remained stubbornly elusive. Herein, we set out to resolve this enigma by employing a hypothesis-driven approach. First, we searched for variants in strong linkage disequilibrium (LD) with rs17107315:T>C using HaploReg v4.1. Second, we identified two candidate SNPs by visual inspection of sequences spanning all 25 SNPs found to be in LD with rs17107315:T>C, guided by prior knowledge of pancreas-specific transcription factors and their cognate binding sites. Third, employing a novel cis-regulatory module (CRM)-guided approach to further filter the two candidate SNPs yielded a solitary candidate causal variant. Finally, combining data from phylogenetic conservation and chromatin accessibility, cotransfection transactivation experiments, and population genetic studies, we suggest that rs142703147:C>A, which disrupts a PTF1L-binding site within an evolutionarily conserved HNF1A-PTF1L CRM located ∼4 kb upstream of the SPINK1 promoter, contributes to the aforementioned chronic pancreatitis risk haplotype. Further studies are required not only to improve the characterization of this functional SNP but also to identify other functional components that might contribute to this high-risk haplotype.

SPINK1 , PRSS1 , CTRC , and CFTR Genotypes Influence Disease Onset and Clinical Outcomes in Chronic Pancreatitis

“…SPINK1 p.N34S is the variant most frequently associated with ICP in European countries, being present in 382 of 5962 (6.4%) combined Caucasian ICP patient alleles and 83 of 11638 (0.07%) combined control alleles (OR = 9.70) 20 . It is pertinent to point out that SPINK1 p.N34S itself has no functional effect but is in complete (in Caucasians) or high (in Han Chinese) linkage disequilibrium with a functional regulatory single nucleotide polymorphism in an upstream enhancer 44 . In the Han Chinese population, SPINK1 p.N34S was present in only 19 of 1430 (1.33%) ICP patient alleles and 5/2392 (0.21%) control alleles (OR = 6.4; P < 0.001), and was much less common than the most frequent variant, SPINK1 c.194+2T>C, whose allele frequency was 350/1430 (24.5%) in ICP patients and 13/2392 (0.54%) in controls (OR = 59.3; Table 2 ).…”

Severe infantile isolated exocrine pancreatic insufficiency caused by the complete functional loss of theSPINK1gene

“…The next most deleterious SPINK1 genotypes, which include those heterozygous SPINK1 variants that render the affected alleles completely nonfunctional (such as large deletions and frameshifting variants), have also been reported to cause chronic pancreatitis (Le Maréchal et al., ; Masson et al., ). By contrast, less deleterious SPINK1 genotypes exemplified by the haplotype harboring the SPINK1 c.101A > G (p.Asn34Ser) variant are generally considered to represent predisposing rather than a causative factors for chronic pancreatitis (Boulling et al., ). In short, SPINK1 genotypes that do not result in a complete functional loss of the SPINK1 gene tend either to predispose or to cause chronic pancreatitis.…”

Toward a clinical diagnostic pipeline for SPINK1 intronic variants

“…It should however be noted that whereas the SPINK1 gene comprises five exons, in accordance with mRNA reference sequence accession NM_003122.3, the gene expressed in the exocrine pancreas comprises only four exons [55, 56]. It is the latter gene structure that is used by both pancreatitis genetics researchers [6, 9, 13, 40, 57] and Ensembl (refer to ENSG00000164266) [58]. The traditional IVS (InterVening Sequence; i.e., an intron) nomenclature for describing SPINK1 intronic variants corresponds to the four-exon gene structure.…”

“…The clinical relevance of canonical splice site variants, nonsense mutations, or large-scale genomic deletions in the SPINK1 gene is generally clear. By contrast, the clinical relevance of SPINK1 promoter and enhancer variants [7–9], missense variants [10, 11], or intronic variants occurring outwith the canonical splice sites [12, 13] has often had to be ascertained by in vitro functional analysis.…”