Genes Encoding Telomere-Binding Proteins TERF1, TERF2 and TIN2 Are mutated in Patients with Acquired Aplastic Anemia.
Authors: Calado Rodrigo T. , Savage Sharon A. , Lansdorp Peter M. , Chanock Stephen J. , Young Neal S. 
Published: October 2019

Abstract
Telomere shortening is observed in some patients with acquired aplastic anemia (AA) and associates with poor response to immunosuppressive therapy. Mutations in components of the telomerase complex (RNA component, TERC, and reverse transcriptase, TERT) have been found in AA patients. As the proportion of AA patients with short telomeres is greater than the number of patients with telomerase complex component mutations, we investigated whether genetic variations in the telomere-binding proteins telomeric repeat binding factor 1 (TERF1), telomeric repeat binding factor 2 (TERF2) and TERF1-interacting nuclear factor 2 (TIN2) were also associated with AA. TERF1 is a negative regulator of telomere length; TIN2 regulates TERF1 function, and TERF2 protects telomeres from degradation and fusion. Bi-directional sequence analysis was performed across all exons and proximal promoter regions of TERF1, TERF2, and TINF2 genes in 147 patients with AA and 118 healthy subjects. Haplotypes were inferred by Phase 2.0. Telomere length of leukocytes was measured by flow cytometry fluorescent in situ hybridization (flow-FISH). A nonsynonymous mutation in TERF1, exon 9, codon 377 (Ala → Val) was found in a 15-year old African-American girl who did not respond to immunosuppressive therapy, but not in controls. A specific TERF1 haplotype was more common in patients (P=0.004), whereas another haplotype was more common in controls (P=0.02). For TERF2 , a mutation in exon 6, codon 273 (Ala→Ser) was found in a 40 year-old Caucasian female patient, but not in controls. A G/A single nucleotide polymorphism (SNP) in exon 6 was present in 15% of patients and 9% of controls (P=0.04). TINF2 was the most polymorphic gene in both patients and contorls. As these variants were found mainly in African-Americans, additional 94 healthy African-American controls were sequenced. In the proximal promoter region, two SNPs and one 16-base-pair deletion were found in patients, but not in 212 controls. The 16-base-pair deletion (−10 to −25) was found in a 31-year-old African-American female who did not respond to immunosuppression, with very short telomeres of leukocytes (Δ TRF −2.59 kb). Also, a -97G/C transition was found in a 66-year-old African-American female who partially responded to immunosupression, also tracking to short telomeres (ΔTRF, −1.22 kb). The −260G/C promoter SNP was present in 3.5% of controls and 0.4% of patients P<0.0001), and the −91C/T SNP was seen in 1.7% of controls but not in patients (P<0.0001). The most common haplotype was seen in 94% of patients and 86% of controls (P=0.003), suggesting an at risk haplotype. A rare haplotype in patients (0.4%) was more common in controls, 3.4% (P=0.01) suggesting a rare but potentially protective variant. These data suggest that mutations and specific haplotypes may confer risk for AA. In addition, we have identified novel mutations in three genes that are essential for telomere stability. In conclusion, mutations in different genes involved in telomere stability and repair appear to be hematologic genetic risk factors for bone marrow failure in adults.