Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.497495
Title: Regulation of the SRF cofactor MAL by actin
Author: Guettler, Sebastian
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2007
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Abstract:
Serum Response Factor (SRF) is controlled by actin dynamics at many of its target genes: Rho-induced depletion of G-actin is sensed by MAL, a member of the myocardin family of SRF coactivators. MAL binds G-actin via its N-terminus, the "RPEL domain", containing three RPEL motifs. MAL rapidly circulates between nucleus and cytoplasm in resting NIH3T3 fibroblasts. It accumulates in the nucleus and activates SRF upon serum stimulation, which alters interactions between G-actin and the RPEL domain. In contrast, myocardin (MC) itself is constitutively nuclear and active when expressed in fibroblasts, suggesting that it is not controlled through Rho. This thesis addresses the mode and functions of actin binding by myocardin-family proteins. Actin binding targets MAL for efficient CRM1-mediated nuclear export. Nuclear accumulation of MAL is not sufficient for activation of SRF-mediated transcription unless an inhibitory MAL-actin interaction in the nucleus is released. Actin therefore fulfils a dual role in MAL regulation by controlling MAL localisation as well as activity. The MAL RPEL domain is sufficient to confer actin-regulated nucleocytoplasmic trafficking and binds multiple actin molecules, efficiently sequestering them from polymerisation. Actin-binding toxins directly interfere with the MAL-actin complex. The RPEL motif represents an actin-binding unit: affinities of MAL RPEL motifs 1 and 2 for actin are relatively high while RPEL3 binds actin weakly. RPEL motifs cooperate to regulate MAL. The regulatory contribution of an RPEL3-actin interaction depends on actin binding by the RPEL 1-2 unit, differences in which account for differential regulation of MAL and MC, which binds actin weakly. A model of MAL regulation by differential actin occupancy of multiple RPEL motifs is proposed. Crystal structures of MAL RPEL motifs 1 and 2 bound to G-actin were obtained. RPEL motifs maintain hydrophobic interactions with a hydrophobic cleft at the subdomain 1-3 interface of actin and a "platform" on subdomain 3, both at the "base" of the actin molecule in its conventional view (Kabsch et al., 1990). The RPEL motif also establishes critical polar interactions with actin. Conservation of the RPEL motif reflects actin binding. The structures rationalise RPEL-actin affinities and competition of actin-binding toxins and profilin with MAL. A crystal structure of the MAL RPEL domain bound to three actin molecules revealed an additional actin-binding site within the RPEL 1-2 linker and actin-actin contacts in the RPEL domain-actin complex.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.497495  DOI: Not available
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