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D.M. Gianferante et al.
Figure 3. Cases of RPL35A large deletions ordered by size in megabases (mB). Each horizontal bar represents the starting and stopping genomic co-ordinates of a RPL35A large deletion case. For literature cases without a case identification (ID) in article, the PubMed ID (PMID) was used as case ID (Online Supplementary Table S3). Thin black vertical lines indicate genomic start- ing and stopping co-ordinates of 3q29 deletion syndrome; thick black verti- cal line is the estimated location of RPL35A. See Online Supplementary Table S5 for exact genes deleted per case.
quency of large deletions in published cases of DBA caused by pathogenic variants in RPL35A is unknown due to the rarity of the disease. However, our findings are con- sistent with what is reported in HGMD where 40% (8 of 21) of DBA cases caused by RPL35A are reported as large/gross deletions.28 Fifty-six percent of non-large dele- tion pathogenic variants were comprised of five variants. Domain information is not well described for this gene, but we did identify apparent clustering of cases between codon 28 to 33 that may be indicative of a potentially important domain (Figure 2). A sufficient number of vari- ants were not available for a variant cluster-phenotype analysis.
In our genotype-phenotype analysis comparing RPL35A large deletions with all other pathogenic variants in RPL35A, patients with large deletions exhibited more severe phenotypes across multiple phenotypic domains, including severity of anemia (steroid-resistance and/or RBC transfusion-dependence), neutropenia, craniofacial abnormalities, chronic GI problems, and intellectual dis- abilities. This suggests that a complex phenotype may be associated with the entire RPL35A gene deletion itself or due to contiguous genes deleted in the 3q29 region. Although not well described in DBA cases caused by RPL35A variants, similar phenotypes in DBA patients with large deletions in other ribosomal genes have been report- ed, including multiple congenital malformations, steroid resistant anemia, and neurodevelopmental abnormalities (e.g., RPL11, RPS17, RPS19, RPS26).18,43 Studies comparing cases with RPL35A large deletion with DBA due to large deletions involving other ribosomal protein genes may elucidate genotype-phenotype relationships and influ- ences of contiguous gene deletions on DBA phenotypes.
Notably, immunodeficiency and recurrent infections were both common features in cases with RPL35A large dele- tions compared with all other pathogenic variants in RPL35A (P=0.01 and 0.03, respectively). Although the number of cases is small, neutropenia appears to be con- tributing to immunodeficiency and recurrent infections, since all these cases were also severely neutropenic and received GCSF treatment. It is unclear from our study whether the severe neutropenia and immunodeficiency seen here is driven by RPL35A gene deletion with or with- out the contribution of contiguous genes deleted in the region or by other LOF variants within RPL35A itself, since severe neutropenia (accompanied by immunodefi- ciency in one case) was also seen in two of the six cases with other LOF variants.
Some of the clinical phenotypes of patients with DBA due to large deletions involving RPL35A are similar to those observed in 3q29 deletion syndrome, including con- genital abnormalities and intellectual disability, and may be influenced by additional genes deleted in the 3q29 region.22,23 Previous speculations suggested genes of impor- tance in 3q29 deletion syndrome studies include PAK2, DLG1, and IL1RAP in intellectual disabilities and TP63 and OSTN in bone defects.24,44-46 However, immunodeficiency and steroid-resistant anemia are not features found in 3q29 deletion syndrome,22,23 making it less likely that a gene deleted in the 3q29 deletion syndrome region is the etiology of the immune and severe hematologic pheno- type seen in our cohort. A case report of DBA caused by a RPL35A large deletion and immune deficiency speculated that RNF168, an autosomal recessive gene located in the 3q29 deletion syndrome region that underlies the immune deficiency Riddle Syndrome, could be a possible candidate
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