A Mild Case of Autosomal Recessive Osteopetrosis Masquerading as the Dominant Form Involving Homozygous Deep Intronic Variations in the CLCN7 Gene

Published online: 26 May 2022Osteopetrosis is a heterogeneous group of rare hereditary diseases characterized by increased bone mass of poor quality. Autosomal-dominant osteopetrosis type II (ADOII) is most often caused by mutation of the CLCN7 gene leading to impaired bone resorption. Autosomal recessive osteopetrosis (ARO) is a more severe form and is frequently accompanied by additional morbidities. We report an adult male presenting with classical clinical and radiological features of ADOII. Genetic analyses showed no amino-acid-converting mutation in CLCN7 but an apparent haploinsufficiency and suppression of CLCN7 mRNA levels in peripheral blood mononuclear cells. Next generation sequencing revealed low-frequency intronic homozygous variations in CLCN7, suggesting recessive inheritance. In silico analysis of an intronic duplication c.595-120_595-86dup revealed additional binding sites for Serine- and Arginine-rich Splicing Factors (SRSF), which is predicted to impair CLCN7 expression. Quantitative backscattered electron imaging and histomorphometric analyses revealed bone tissue and material abnormalities. Giant osteoclasts were present and additionally to lamellar bone, and abundant woven bone and mineralized cartilage were observed, together with increased frequency and thickness of cement lines. Bone mineralization density distribution (BMDD) analysis revealed markedly increased average mineral content of the dense bone (CaMean T-score + 10.1) and frequency of bone with highest mineral content (CaHigh T-score + 19.6), suggesting continued mineral accumulation and lack of bone remodelling. Osteocyte lacunae sections (OLS) characteristics were unremarkable except for an unusually circular shape. Together, our findings suggest that the reduced expression of CLCN7 mRNA in osteoclasts, and possibly also osteocytes, causes poorly remodelled bone with abnormal bone matrix with high mineral content. This together with the lack of adequate bone repair mechanisms makes the material brittle and prone to fracture. While the skeletal phenotype and medical history were suggestive of ADOII, genetic analysis revealed that this is a possible mild case of ARO due to deep intronic mutation.Jochen G. Hofstaetter, Gerald J. Atkins, Hajime Kato, Masakazu Kogawa, Stéphane Blouin, Barbara M. Misof, Paul Roschger, Andreas Evdokiou, Dongqing Yang, Lucian B. Solomon, David M. Findlay, Nobuaki It

RNA in phylogenetic reconstruction

Ribosomal RNA (rRNA) genes are the most widely used data source in phylogenetic reconstruction. They are highlystructured, with large parts of the molecules exhibiting very strong conservation of their base pairing patterns. Since also sequence conservation varies dramatically between different regions of rRNA genes, these data are informative on a wide range of phylogenetic timescales, ranging from recent to ancient splits. But individual columns of rRNA alignments are not independent. This is a consequence of biologically functional secondary structures with highly varying degrees of conservation. Therefore, any correlations within sequence alignments of structurally functional RNA will distort the phylogenetic signal and/or lead to gross overestimates of tree stability. On the other hand, alignment accuracy can be improved substantially by incorporating secondary structure conservation. Maximum Likelihood and Bayesian approaches are amenable to using RNA-specific sub stitution models that treat conserved base pairs appropriately, but they require accurate secondary structure models as input. Structure prediction algorithms for single RNA sequences are well known and widely used; but it is not straightforward to apply these predictions for phylogenetic purposes. The main limitation is that accuracy of thermodynamic folding algorithms declines sharply as the length of the RNA increases, this is in part due toinaccuracies within the thermodynamic folding parameters, in part caused by the kinetics of folding process and tertiary interactions, and even more in the fact that RNA and protein components of the ribosome are tightly packed and thus mutually influence their folds. The functional rRNA structures, therefore, cannot reasonably be expected to be identical with the minimum free energy structures of isolated rRNAs computed by thermodynamic folding algorithms