Newly synthetized histone H3 is degraded by chaperone mediated autophagy

Hormazabal , Juan.; Saavedra, Francisco.; Loyola, Alejandra.; Alfaro, Ivan E.


Introduction: Histones are proteins involved in DNA compaction. Biogenesis, transport, and assembly of histones are highly regulated processes assisted by chaperones. Little is known about the proteostatic mechanisms of recently synthesized histones in mammals. Hsc70 is a chaperone that regulates cytosolic histone biogenesis and is also involved in the recognition of proteins selectively targeted to degradation by chaperone-mediated autophagy (CMA). We identified two KFERQ-like CMA-targeting motifs in the sequence of H3. However, there are no direct evidences of CMA involvement in the control of histone protein levels. Materials and Methods: We used lysosomal fractions isolated from rat livers to determinate the uptake of histone H3 to lysosomes in vitro. We also used cytosolic protein extracts from HeLa cells to study H3 degradation in pulse and chase experiments. These cells were also used to study transport of fluorescent tagged H3 to lysosomes by imaging microscopy in control and Lamp2a knockdown cells. Results: We demonstrated the presence of H3 in lysosomes isolated from rat liver and a fraction of cytosolic H3 degraded by lysosomes. Purified recombinant H3 is transported into isolated lysosomes confirming that H3 is a bona fide CMA substrate. Moreover, fluorescent-tagged histones associate with the lysosomal compartment in HeLa cells in response to nutrient starvation and knockdown of Lamp2a in HeLa cells increases the levels of cytosolic H3. Interestingly, NASP, a protein involved in stabilization of H3 cytosolic pool, is able to displace H3 from Hsc70 binding in vitro and protects H3 from degradation in isolated lysosomes, thus linking histone biogenesis processes to CMA mediated H3 degradation. Discussion: Our results suggest a critical role of CMA in the control of cytosolic histone levels, a process that would be connected to the mechanism of histone biogenesis. Acknowledgements: Fondecyt 11161056, Fondecyt 1160480, Proyecto FCPV Apoyo Basal AFB170004

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Fecha de publicación: 2018
Año de Inicio/Término: 18 Octubre de 2018