Fare Miyoblast Hücre Hattında KLF5 Geninin Farklılaşma Sürecine Etkilerinin Araştırılması

Abstract

Embryonic development of the skeletal muscle and injury repair share several molecular events such as the myogenic regulatory factors (MRFs) and paired-homeobox transcription factors. Myogenesis is a highly regulated complex process and several other regulatory proteins and mechanisms that control activation, proliferation, commitment, migration and fusion of the myoblasts are studying hardly. Our previous studies shown a zinc finger protein, Klf5 interplays in muscle differentiation. Basal expression of Klf5 in myoblasts is upregulated in the course of differentiation and myotube formation in vitro. Similarly, Klf5 is also upregulated in vivo during acute injury regeneration concomitantly with the formation of young myofibers and returns to the baseline upon restoration of the tissue architecture. Likewise, Klf5 expression is also induced in chronic degenerative conditions such as the inherited myopathies and representative animal models. Klf5 was sumoylated and translocated to the nucleus with differentiation in vitro and regeneration in vivo. Experimental studies that are conceived for this thesis work that included shRNA mediated conditional silencing Klf5 showed that Klf5 regulated the proliferation rate of myoblasts but as well as the fusion and myotube maturation stages of differentiation. On the other hand, overexpression of Klf5 was shown to induce cell cycle arrest. These findings are also confirmed by the disruption of Klf5 function using a dominant negative Klf5 isoform. The expression analysis of myogenic differentiation markers such as M-cadherin, NCAM, Desmin as well as DNA quantitation based proliferation rate analysis of myoblasts further supported these findings. In vitro studies also revealed that Klf5 also exerted a role in motility of the myoblasts beside proliferation and differentiation. Studies were conducted to reveal the plausible interactions between Klf5 and other MRFs. Results suggest that Klf5 implies a regulatory function over Myogenin. However, the master regulator MyoD is apparently counteracting the inhibitor functions of Klf5. This data provide evidence that Klf5 is a novel modulator of the proliferation and differentiation of myoblasts, impacting skeletal muscle regeneration.