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Buson, Lisa (2019) Identification of specific non-coding RNAs involved in skeletal muscle metabolism: a single cell approach. [Ph.D. thesis]

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Abstract (italian or english)

Skeletal muscle is a heterogeneous tissue composed of different cell types with myofibers that are the smallest complete contractile system that influence muscle contraction velocity and metabolism. Myofibers, can be classified in three main categories: slow oxidative (also known as type I), fast-oxidative (or type IIA) and fast-glycolytic (or type IIB).
Recently, the non-coding RNAs have emerged as a new RNA class with regulatory functions in several biological processes. Non-coding RNAs (ncRNAs) are transcripts with little or no coding capacity, divisible in two classes: microRNAs (miRNAs; shorter than 200nt) and long non-coding RNAs (lncRNAs; longer than 200nt).
To uncover the functions of ncRNAs in muscle physiopathology, we profiled both miRNAs and lncRNAs in single myofibers, evidencing the importance of single cell approaches.
miRNAs. We discovered two circuits that connect myofiber metabolic traits with mir-27a-3p and mir-142-3p. We evidenced that miR-27a-3p was highly expressed in oxidative myofibers while miR-142-3p was low expressed, suggesting a complementary mechanism in the regulation of myofiber metabolism. In fact, we demonstrated a) the involvement of miR-27a-3p in the inhibition of the synthesis of Phosphoglucomutase 2 (Pgm2) and acid α-glucosidase (Gaa) enzymes, both involved in glycogenolysis and b) the control of the Fndc5-Irisin pathway by miR-142-3p that instead modulate lipids content within myofibers.
lncRNAs. Since there are no data available on lncRNAs expressed by myofibers, we defined them according to their expression in skeletal muscle fibers. We showed that lncRNAs are fiber type specific, with a peculiar subcellular localization and are sensible to muscle atrophy. We showed that the lncRNA Plasmacytoma variant translocation 1 (Pvt1) is activated early during muscle atrophy and it is more expressed in fast myofibers. These observations were the starting points for the pathway dissection throughout which Pvt1 acts in skeletal muscle. In fact, atrophy causes metabolic changes and myofiber switching, both inter-related myofiber characteristics. We demonstrated the impact of Pvt1 on mitochondrial respiration and morphology, its regulatory activity on autophagy and apoptosis, and its consequent ability to influence myofiber size. Pvt1 regulates mitochondria physiology though its capacity in modulating c-Myc, which in turn, regulates Bcl2, Bax/Bak, Mfn1, and Becn1, impinging mitochondrial dynamics and finally muscle fiber dimension.
The story is then made difficult by the fact that Pvt1 locus codify also for miRNAs. We showed that three miRNAs encoded by Pvt1 locus (miR-1207-3p, miR-1207-5p and miR-1208) seems influence mitochondrial dynamics. We speculate that these miRNAs could target the lncRNA Pvt1 revealing a new regulatory loop of miRNAs and lncRNAs that may govern skeletal muscle homeostasis.
In skeletal muscle research, the evolution of new single cell approaches allows the identification of signatures that specify functional and metabolic behavior of myofibers, otherwise impossible to detect analyzing the whole muscle.
Our works contribute to better explain not only the importance of single cell studies, but also how ncRNAs participate in the regulation of skeletal muscle functions.

EPrint type:Ph.D. thesis
Tutor:Lanfranchi, Gerolamo and Cagnin, Stefano
Supervisor:Bubacco, Luigi and DePitta', Cristiano
Data di deposito della tesi:01 December 2019
Anno di Pubblicazione:01 December 2019
Key Words:Non-coding RNAs Skeletal muscle Single cell approach
Settori scientifico-disciplinari MIUR:Area 05 - Scienze biologiche > BIO/11 Biologia molecolare
Struttura di riferimento:Centri > Centro Interdipartimentale di servizi A. Vallisneri
Dipartimenti > Dipartimento di Biologia
Codice ID:12240
Depositato il:02 Feb 2021 11:32
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