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Summary: Osteoblasts orchestrate bone formation through the secretion of type I collagen and other constituents of the matrix on which hydroxyapatite crystals mineralize. Here, we show that TENT5A, whose mutations were found in congenital bone disease osteogenesis imperfecta patients, is a cytoplasmic poly(A) polymerase playing a crucial role in regulating bone mineralization. Direct RNA sequencing revealed that TENT5A is induced during osteoblast differentiation and polyadenylates mRNAs encoding Col1α1, Col1α2, and other secreted proteins involved in osteogenesis, increasing their expression. We postulate that TENT5A, possibly together with its paralog TENT5C, is responsible for the wave of cytoplasmic polyadenylation of mRNAs encoding secreted proteins occurring during bone mineralization. Importantly, the Tent5a knockout (KO) mouse line displays bone fragility and skeletal hypomineralization phenotype resulting from quantitative and qualitative collagen defects. Thus, we report a biologically relevant posttranscriptional regulator of collagen production and, more generally, bone formation.

Osteoblasts orchestrate bone formation by secreting dense, highly cross-linked type I collagen and other proteins involved in osteogenesis. Mutations in Col1α1, Col1α2, or collagen biogenesis factors lead to the human genetic disease, osteogenesis imperfecta (OI). Herein, we show that the TENT5A gene, whose mutation is responsible for poorly characterized type XVIII OI, encodes an active cytoplasmic poly(A) polymerase regulating osteogenesis. TENT5A is induced during osteoblast differentiation and TENT5A KO osteoblasts are defective in mineralization. The TENT5A KO mouse recapitulates OI disease symptoms such as bone fragility and hypomineralization. Direct RNA sequencing revealed that TENT5A polyadenylates and increases expression of Col1α1 and Col1α2 RNAs, as well as those of other genes mutated in OI, resulting in lower production and improper folding of collagen chains. Thus, we have identified the specific pathomechanism of XVIII OI and report for the first time a biologically relevant post-transcriptional regulator of collagen production. We further postulate that TENT5A, possibly together with its paralogue TENT5C, is responsible for the wave of cytoplasmic polyadenylation of mRNAs encoding secreted proteins occurring during bone mineralization.

TENT5C is a non-canonical cytoplasmic poly(A) polymerase highly expressed by activated B cells to suppress their proliferation. Here we measure the global distribution of poly(A) tail lengths in responsive B cells using a Nanopore direct RNA-sequencing approach, showing that TENT5C polyadenylates immunoglobulin mRNAs regulating their half-life and consequently steady-state levels. TENT5C is upregulated in differentiating plasma cells by innate signaling. Compared with wild-type, Tent5c−/− mice produce fewer antibodies and have diminished T-cell-independent immune response despite having more CD138high plasma cells as a consequence of accelerated differentiation. B cells from Tent5c−/− mice also have impaired capacity of the secretory pathway, with reduced ER volume and unfolded protein response. Importantly, these functions of TENT5C are dependent on its enzymatic activity as catalytic mutation knock-in mice display the same defect as Tent5c−/−. These findings define the role of the TENT5C enzyme in the humoral immune response., Regulating polyadenylation is important for mRNA stability, which can in turn affect B cell maturation and humoral immune responses. Here the authors use Nanopore poly(A) sequencing to explore the importance of the cytoplasmic poly(A) polymerase TENT5C, particularly in the production of immunoglobulins.

SummaryTENT5C is a non-canonical cytoplasmic poly(A) polymerase (ncPAP) upregulated in activated B cells and suppressing their proliferation. Herein we measured the global distribution of poly(A) tail lengths in responsive B cells using a modified Nanopore direct RNA-sequencing approach and revealed that TENT5C polyadenylates immunoglobulin mRNAs regulating their steady-state levels. Consequently, TENT5C deficient B cells secrete less antibodies and KO mice have diminished gamma globulin concentrations despite the increased number of CD138high plasma cells as a consequence of accelerated differentiation. TENT5C is explicitly upregulated in differentiating plasma cells by innate signaling. Importantly, TENT5C deficiency in B lymphocytes impairs the capacity of the secretory pathway through the reduction of ER volume and downregulation of unfolded protein response.Our findings define the role of the TENT5C enzyme in B cell physiology and discover the first ncPAP engaged in the regulation of immunoglobulin mRNA poly(A) tails, thus serving as a regulator of humoral immunity.