Biology subjects

Sparago, E.

Publications and source records attributed to Sparago, E..

3 recordsLinked to original sources

bioRxiv2026

Decreased tRNA abundance contributes to decreased translation elongation rate in a prolonged mitosis

Cells undergo dramatic structural rearrangements upon entering mitosis. In addition, the biochemistry of mitotic cells is dramatically altered, including a significant decrease in protein synthesis. The majority of studies of mitotic translation have used cells synchronized by cell cycle altering drugs in transformed cells and much less is known about mitotic translation in primary cells under native conditions. Previous work has found that mitosis activates the integrated stress response (ISR) to trigger eIF2 phosphorylation, but little is known about the input for this response. In this study, we focus on mitotic translational regulation in an immortalized, non-transformed cell line. We confirm decreased mitotic protein synthesis in primary cells and under native conditions. Additionally, we confirm activation of the ISR by phosphorylation of eIF2 during both normal and prolonged mitosis. Interestingly, we also find that decreased translational elongation during mitosis, as evidenced by increased eEF2 phosphorylation and a slower elongation rate. Analysis of mitotic ribosome profiling data revealed an increase in pausing at Alanine-GCG codons during mitosis and a decreased abundance of its cognate tRNA-AlaCGC by northern blotting. Decreased tRNA-AlaCGC is likely sustained by the inability to synthesize additional tRNA due to RNAPol III inhibition in mitosis, yielding an stronger effect with an increased time in mitosis. These results suggest that decreased translation elongation in mitosis triggers inhibition of initiation to decrease global protein synthesis.

cell biology↗

bioRxiv2025

Role of the SAF-A/HNRNPU ATPase and RGG domains in X chromosome inactivation, nuclear dynamics, transcription, splicing, and cell proliferation

The SAF-A/HNRNPU gene encodes an abundant nuclear protein conserved throughout vertebrates, and is mutated in individuals with HNRNPU syndrome, a neurological human disease. SAF-A is important for maintaining lncRNA localization, splicing, and gene expression state. The mechanistic role of SAF-A in each of these processes is coordinated by one or more of its functional domains, which include an N-terminal SAP domain, a central ATPase domain, and a series of C-terminal RGG repeats embedded in a low-complexity region. The SAP domain and RGG repeats define two nucleic acid interaction domains, with both capable of binding DNA or RNA. Here we use an allelic reconstitution strategy to investigate the role of the SAF-A ATPase domain and RGG repeats. We show that both the ATPase and RGG repeats control SAF-A nuclear dynamics, and present genetic evidence that SAF-A interacts with nascent transcripts through the RGG repeats. The SAF-A ATPase domain and RGG repeats were also required for maintaining XIST RNA and facultative heterochromatin marks on the inactive X chromosome, with distinct effects of mutations that block ATP binding and ATP hydrolysis. Analysis of transcriptome datasets revealed that the SAF-A ATPase domain and RGG repeats are both required for proper mRNA splicing, but not for gene expression. Importantly, we found that like the SAP domain, the SAF-A ATPase domain and RGG repeats are required for cell proliferation. Collectively, our findings highlight the importance of the SAF-A SAP, ATPase and RGG domains in essential functions of nuclear biology. These analyses will therefore inform our understanding of the disease state in HNRNPU syndrome.

cell biology↗

bioRxiv2024

Role of the SAF-A SAP domain in X inactivation, transcription, splicing, and cell proliferation

SAF-A is conserved throughout vertebrates and has emerged as an important factor regulating a multitude of nuclear functions, including lncRNA localization, gene expression, and splicing. SAF-A has several functional domains, including an N-terminal SAP domain that binds directly to DNA. Phosphorylation of SAP domain serines S14 and S26 are important for SAF-A localization and function during mitosis, however whether these serines are involved in interphase functions of SAF-A is not known. In this study we tested for the role of the SAP domain, and SAP domain serines S14 and S26 in X chromosome inactivation, protein dynamics, gene expression, splicing, and cell proliferation. Here we show that the SAP domain serines S14 and S26 are required to maintain XIST RNA localization and polycomb-dependent histone modifications on the inactive X chromosome in female cells. In addition, we present evidence that an Xi localization signal resides in the SAP domain. We found that that the SAP domain is not required to maintain gene expression and plays only a minor role in mRNA splicing. In contrast, the SAF-A SAP domain, in particular serines S14 and S26, are required for normal protein dynamics, and to maintain normal cell proliferation. We propose a model whereby dynamic phosphorylation of SAF-A serines S14 and S26 mediates rapid turnover of SAF-A interactions with DNA during interphase.

cell biology↗