Optogenetic Control of Myocardin‐Related Transcription Factor A Subcellular Localization and Transcriptional Activity Steers Membrane Blebbing and Invasive Cancer Cell Motility

Abstract

Similar to SRF, its coactivator MRTF-A (also known as MAL or MKL1) is ubiquitously expressed and controls actin-associated gene expression for numerous functions in development and disease as well as many cell migratory processes. [3,4] The latter occurs largely via the regulation of cytoskeletal genes following signal-induced MRTF-A nuclear translocation. [5][6][7] Notably, MRTF-A dependent immediate gene expression appears to be the predominant cellular SRF transcriptional response to extracellular signals such as serum. [8] In addition, it is now well established that MRTF-A transcriptional activity is an important driver of tumor progression by promoting invasion and metastasis. [9][10][11][12][13][14][15][16] We could recently show that MRTF-A activity promotes non-apoptotic plasma membrane (PM) blebbing often observed during blebassociated invasive cell migration. [17] This MRTF-A nuclear activity was in turn triggered by PM blebbing thereby facilitating a positive feedback loop to drive PM dynamics and cell motility through MRTF-SRF transcription. [17] Such mechanism is consistent with the fact that nuclear localization of MRTF-A depends on actin dynamics. [5] This occurs via three core RPEL actin-binding sites (R1-R3) and the two spacer actin-binding regions (S1, S2) within the N-terminus of MRTF-A (Figure 1A). [18] Specifically, the RPEL domain of MRTF-A interacts with actin such that binding of all five actin monomers (G-actin) promotes MRTF-A nuclear export while dissociation of the G-actin molecules favors its Nuclear localization signal (NLS)-dependent nuclear import (Figure 1A). [18] This mechanism allows MRTF-A to sense and rapidly respond to changes in actin dynamics resulting in the regulation of gene expression. [19] Nucleocytoplasmic shuttling is regarded as an essential mechanism to control the activity of transcriptional factors. [20] Quite recently, a few optogenetic tools have been generated to artificially manipulate the localization of nuclear proteins, [21] providing a new perspective for modulating cellular activities. Here, we devised and employed an optogenetic strategy using the modified light-oxygen-voltage-sensing domain 2 (LOV2) LEXY to spatiotemporally control MRTF-A nuclear localization and function independently of actin dynamics. Optogenetic The myocardin-related transcription factor A (MRTF-A) controls the transcriptional activity of the serum response factor (SRF) in a tightly controlled actin-dependent manner. In turn, MRTF-A is crucial for many actin-dependent processes including adhesion, migration, and contractility and has emerged as a novel target for anti-tumor strategies. MRTF-A rapidly shuttles between cytoplasmic and nuclear compartment via dynamic actin interactions within its N-terminal RPEL domain.Here, optogenetics is used to spatiotemporally control MRTF-A nuclear localization by blue light using the light-oxygen-voltage-sensing domain 2-domain based system LEXY (light-inducible nuclear export system). It is found that lightregulated nuclear export of MRTF-A o...