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Neurons critically rely on the functions of RNA-binding proteins to maintain their polarity and resistance to neurotoxic stress. HnRNP R has a diverse range of post-transcriptional regulatory functions and is important for neuronal development by regulating axon growth. Hnrnpr pre-mRNA undergoes alternative splicing giving rise to a full-length protein and a shorter isoform lacking its N-terminal acidic domain. To investigate functions selectively associated with the full-length hnRNP R isoform, we generated a Hnrnpr knockout mouse (Hnrnpr(tm1a/tm1a)) in which expression of full-length hnRNP R was abolished while production of the truncated hnRNP R isoform was retained. Motoneurons cultured from Hnrnpr(tm1a/tm1a) mice did not show any axonal growth defects but exhibited enhanced accumulation of double-strand breaks and an impaired DNA damage response upon exposure to genotoxic agents. Proteomic analysis of the hnRNP R interactome revealed the multifunctional protein Yb1 as a top interactor. Yb1-depleted motoneurons were defective in DNA damage repair. We show that Yb1 is recruited to chromatin upon DNA damage where it interacts with gamma-H2AX, a mechanism that is dependent on full-length hnRNP R. Our findings thus suggest a novel role of hnRNP R in maintaining genomic integrity and highlight the function of its N-terminal acidic domain in this context.

Gene expression requires tight coordination of the molecular machineries that mediate transcription and splicing. While the interplay between transcription kinetics and spliceosome fidelity has been investigated before, less is known about mechanisms regulating the assembly of the spliceosomal machinery in response to transcription changes. Here, we report an association of the Smn complex, which mediates spliceosomal snRNP biogenesis, with the 7SK complex involved in transcriptional regulation. We found that Smn interacts with the 7SK core components Larp7 and Mepce and specifically associates with 7SK subcomplexes containing hnRNP R. The association between Smn and 7SK complexes is enhanced upon transcriptional inhibition leading to reduced production of snRNPs. Taken together, our findings reveal a functional association of Smn and 7SK complexes that is governed by global changes in transcription. Thus, in addition to its canonical nuclear role in transcriptional regulation, 7SK has cytosolic functions in fine-tuning spliceosome production according to transcriptional demand.

Neurons critically rely on the functions of RNA-binding proteins to maintain their polarity and resistance to neurotoxic stress. HnRNP R has a diverse range of post-transcriptional regulatory functions and is important for neuronal development by regulating axon growth. Hnrnpr pre-mRNA undergoes alternative splicing giving rise to a full-length protein and a shorter isoform lacking its N-terminal acidic domain. To investigate functions selectively associated with the full-length hnRNP R isoform, we generated a Hnrnpr knockout mouse (Hnrnpr(tm1a/tm1a)) in which expression of full-length hnRNP R was abolished while production of the truncated hnRNP R isoform was retained. Motoneurons cultured from Hnrnpr(tm1a/tm1a) mice did not show any axonal growth defects but exhibited enhanced accumulation of double-strand breaks and an impaired DNA damage response upon exposure to genotoxic agents. Proteomic analysis of the hnRNP R interactome revealed the multifunctional protein Yb1 as a top interactor. Yb1-depleted motoneurons were defective in DNA damage repair. We show that Yb1 is recruited to chromatin upon DNA damage where it interacts with gamma-H2AX, a mechanism that is dependent on full-length hnRNP R. Our findings thus suggest a novel role of hnRNP R in maintaining genomic integrity and highlight the function of its N-terminal acidic domain in this context.

Gene expression requires tight coordination of the molecular machineries that mediate transcription and splicing. While the interplay between transcription kinetics and spliceosome fidelity has been investigated before, less is known about mechanisms regulating the assembly of the spliceosomal machinery in response to transcription changes. Here, we report an association of the Smn complex, which mediates spliceosomal snRNP biogenesis, with the 7SK complex involved in transcriptional regulation. We found that Smn interacts with the 7SK core components Larp7 and Mepce and specifically associates with 7SK subcomplexes containing hnRNP R. The association between Smn and 7SK complexes is enhanced upon transcriptional inhibition leading to reduced production of snRNPs. Taken together, our findings reveal a functional association of Smn and 7SK complexes that is governed by global changes in transcription. Thus, in addition to its canonical nuclear role in transcriptional regulation, 7SK has cytosolic functions in fine-tuning spliceosome production according to transcriptional demand.

Neurotransmission defects and motoneuron degeneration are hallmarks of spinal muscular atrophy, a monogenetic disease caused by the deficiency of the SMN protein. In the present study, we show that systemic application of R-Roscovitine, a Cav2.1/Cav2.2 channel modifier and a cyclin-dependent kinase 5 (Cdk-5) inhibitor, significantly improved survival of SMA mice. In addition, R-Roscovitine increased Cav2.1 channel density and sizes of the motor endplates. In vitro, R-Roscovitine restored axon lengths and growth cone sizes of Smn-deficient motoneurons corresponding to enhanced spontaneous Ca2+ influx and elevated Cav2.2 channel cluster formations independent of its capability to inhibit Cdk-5. Acute application of R-Roscovitine at the neuromuscular junction significantly increased evoked neurotransmitter release, increased the frequency of spontaneous miniature potentials, and lowered the activation threshold of silent terminals. These data indicate that R-Roscovitine improves Ca2+ signaling and Ca2+ homeostasis in Smn-deficient motoneurons, which is generally crucial for motoneuron differentiation, maturation, and function.

Neurotransmission defects and motoneuron degeneration are hallmarks of spinal muscular atrophy, a monogenetic disease caused by the deficiency of the SMN protein. In the present study, we show that systemic application of R-Roscovitine, a Cav2.1/Cav2.2 channel modifier and a cyclin-dependent kinase 5 (Cdk-5) inhibitor, significantly improved survival of SMA mice. In addition, R-Roscovitine increased Cav2.1 channel density and sizes of the motor endplates. In vitro, R-Roscovitine restored axon lengths and growth cone sizes of Smn-deficient motoneurons corresponding to enhanced spontaneous Ca2+ influx and elevated Cav2.2 channel cluster formations independent of its capability to inhibit Cdk-5. Acute application of R-Roscovitine at the neuromuscular junction significantly increased evoked neurotransmitter release, increased the frequency of spontaneous miniature potentials, and lowered the activation threshold of silent terminals. These data indicate that R-Roscovitine improves Ca2+ signaling and Ca2+ homeostasis in Smn-deficient motoneurons, which is generally crucial for motoneuron differentiation, maturation, and function.

Neurotransmission defects and motoneuron degeneration are hallmarks of spinal muscular atrophy, a monogenetic disease caused by the deficiency of the SMN protein. In the present study, we show that systemic application of R-Roscovitine, a Cav2.1/Cav2.2 channel modifier and a cyclin-dependent kinase 5 (Cdk-5) inhibitor, significantly improved survival of SMA mice. In addition, R-Roscovitine increased Cav2.1 channel density and sizes of the motor endplates. In vitro, R-Roscovitine restored axon lengths and growth cone sizes of Smn-deficient motoneurons corresponding to enhanced spontaneous Ca2+ influx and elevated Cav2.2 channel cluster formations independent of its capability to inhibit Cdk-5. Acute application of R-Roscovitine at the neuromuscular junction significantly increased evoked neurotransmitter release, increased the frequency of spontaneous miniature potentials, and lowered the activation threshold of silent terminals. These data indicate that R-Roscovitine improves Ca2+ signaling and Ca2+ homeostasis in Smn-deficient motoneurons, which is generally crucial for motoneuron differentiation, maturation, and function.

Neurotransmission defects and motoneuron degeneration are hallmarks of spinal muscular atrophy, a monogenetic disease caused by the deficiency of the SMN protein. In the present study, we show that systemic application of R-Roscovitine, a Cav2.1/Cav2.2 channel modifier and a cyclin-dependent kinase 5 (Cdk-5) inhibitor, significantly improved survival of SMA mice. In addition, R-Roscovitine increased Cav2.1 channel density and sizes of the motor endplates. In vitro, R-Roscovitine restored axon lengths and growth cone sizes of Smn-deficient motoneurons corresponding to enhanced spontaneous Ca2+ influx and elevated Cav2.2 channel cluster formations independent of its capability to inhibit Cdk-5. Acute application of R-Roscovitine at the neuromuscular junction significantly increased evoked neurotransmitter release, increased the frequency of spontaneous miniature potentials, and lowered the activation threshold of silent terminals. These data indicate that R-Roscovitine improves Ca2+ signaling and Ca2+ homeostasis in Smn-deficient motoneurons, which is generally crucial for motoneuron differentiation, maturation, and function.