β- and γ-Actins in the nucleus of human melanoma A375 cells (original) (raw)
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New Insights into Cellular Functions of Nuclear Actin
Biology
Actin is one of the most abundant proteins in eukaryotic cells. There are different pools of nuclear actin often undetectable by conventional staining and commercial antibodies used to identify cytoplasmic actin. With the development of more sophisticated imaging and analytical techniques, it became clear that nuclear actin plays a crucial role in shaping the chromatin, genomic, and epigenetic landscape, transcriptional regulation, and DNA repair. This multifaceted role of nuclear actin is not only important for the function of the individual cell but also for the establishment of cell fate, and tissue and organ differentiation during development. Moreover, the changes in the nuclear, chromatin, and genomic architecture are preamble to various diseases. Here, we discuss some of the newly described functions of nuclear actin.
Nuclear actin dynamics – From form to function
FEBS Letters, 2008
Cell biological functions of actin have recently expanded from cytoplasm to nucleus, with actin implicated in such diverse processes as gene expression, transcription factor regulation and intranuclear motility. Actin in the nucleus seems to behave differently than in the cytoplasm, raising new questions regarding the molecular mechanisms by which actin functions in cells. In this review, I will discuss dynamic properties of nuclear actin that are related to its polymerization cycle and nucleocytoplasmic shuttling. By comparing the behaviour of nuclear and cytoplasmic actin and their regulators, I try to dissect the underlying differences of these equally important cellular actin pools.
Conformation-specific antibodies reveal distinct actin structures in the nucleus and the cytoplasm
Journal of Structural Biology, 2005
For many years the existence of actin in the nucleus has been doubted because of the lack of phalloidin staining as well as the failure to document nuclear actin filaments by electron microscopy. More recent findings reveal actin to be a component of chromatin remodeling complexes and of the machinery involved in RNA synthesis and transport. With distinct functions for
Tracking down the different forms of nuclear actin
Trends in Cell Biology
Actin is a rather uncommitted protein with a high degree of structural plasticity: it can adopt a variety of structural states, depending on the specific ionic conditions or the interaction with ligand proteins. These interactions lock actin into a distinct conformation, which specifies the oligomeric or polymeric form it can assume. The interplay between monomeric, oligomeric and polymeric forms is used by the cell to execute an enormous variety of motility processes, such as lamellipodium formation during locomotion or intracellular transport of vesicles. In these cytoplasmic events, monomeric G-actin and filamentous F-actin are the prevalent forms. However, there might be other structural states of actin in cells that have so far not received the attention they deserve. Here, we propose that specific, "unconventional" actin conformations might contribute especially to the multitude of functions executed by actin in the nucleus. We present evidence for the existence of d...
Analysis of nuclear actin-interacting proteins and actin-regulated transcription factors
2019
Actin is best-known from its functions in the cytoplasm, where it is a key component of the cytoskeleton. Cytoskeleton is vital for cells as it enables cell movement and maintains cell shape. Nevertheless, functions of actin are not restricted to the cytoplasm, since actin is also present in the nucleus, where it has been linked to multiple functions from gene activation to chromatin remodeling. Live cell imaging with different nuclear actin probes have demonstrated the importance of actin dynamics inside the nucleus, but the molecular mechanisms by which actin operates in the nucleus are still poorly understood. This is mainly because of the lack of wellcharacterized binding partners for nuclear actin. Therefore, the aim of this thesis was to identify and characterize novel nuclear actin-binding partners and elucidate the molecular mechanisms behind actin regulated transcription factors. To identify nuclear actin-binding partners, we used two complementary mass spectrometry (MS) techniques, affinity purification combined with MS (AP-MS) and proximity dependent biotin identification with MS (BioID). AP-MS protocol was optimized to preserve complete nuclear complexes and BioID was geared towards identifying more transient interactions. We utilized different actin constructs to discriminate nuclear versus cytoplasmic interactions and to assess the requirement for actin polymerization for the putative nuclear interactions. Analysis of our interactome data revealed that actin can form stable complexes with proteins related to chromatin remodeling but seems to function in a dynamic fashion in other nuclear processes, such as transcription and DNA replication. In our experimental setup actin seemed to be monomeric when it associated with nuclear complexes. We also discovered a novel actin-containing complex, human Ada-Two-A-containing complex (hATAC). HATAC is a histone modifying complex and further studies showed that actin directly binds one of it subunits, lysine acetyltransferase 14 (KAT14). We showed that actin-binding modulates histone acetyl transferase (HAT) activity of KAT14 in vitro and in cells. We obtained numerous RNA splicing and mRNA processing factors with our BioID approach, which led us to investigate the role of actin in RNA splicing. Bimolecular fluorescent complementation (BiFC) assays demonstrated that actin associates with different splicing factors and we further showed, for the first time, that actin has a functional role in mRNA splicing, as alterations in nuclear actin levels disturbed survival motor neuron protein 2 (SMN2) alternative exon skipping. In addition, the nuclear actin interactome analysis provided new insights into nuclear processes already earlier linked to actin, such as chromatin remodeling, transcription and DNA replication, and hence this work provides a protein interaction platform for further mechanistic studies of nuclear actin-dependent functions. This thesis work has thereby broadened the knowledge of nuclear actin-binding partners as well as revealed novel regulatory properties of actin-regulated RPEL domain containing proteins.
Differential sublocalization of actin variants within the nucleus
Cytoskeleton, 2010
Conventional actin has been implicated in various nuclear processes including chromatin remodeling, transcription, nuclear transport, and overall nuclear structure. Moreover, actin has been identified as a component of several chromatin remodeling complexes present in the nucleus. In animal cells, nuclear actin exists as a dynamic equilibrium of monomers and polymers. Actin-binding proteins (ABPs) such as ADF/cofilin and profilin play a role in actin import and export, respectively. However, very little is known about the localization and roles of nuclear actin in plants. In multicellular plants and animals, actin is comprised of an ancient and divergent family of protein variants. Here, we have investigated the presence and differential localization of two ancient subclasses of actin in isolated Arabidopsis nuclei. Although the subclass 1 variants ACT2 and ACT8 and subclass 2 variant ACT7 were found distributed throughout the nucleoplasm, ACT7 was often found more concentrated in nuclear speckles than subclass 1 variants. The nuclei from the act2-1/act8-2 double null mutant and the act7-5 null mutant lacked their corresponding actin variants. In addition, serial sectioning of several independent nuclei revealed that ACT7 was notably more abundant in the nucleolus than the subclass 1 actins. Profilin and ADF proteins were also found in significant levels in plant nuclei. The possible functions of differentially localized nuclear actin variants are discussed. V C 2010 Wiley-Liss, Inc.
Embo Journal, 2011
Nuclear actin and actin related proteins (Arps) are integral components of various chromatin remodeling complexes. Actin in such nuclear assemblies does not form filaments but associates in defined complexes, for instance with Arp4 and Arp8 in the INO80 remodeler. To understand the relationship between nuclear actin and its associated Arps and to test the possibility that Arp4 and Arp8 help maintain actin in defined states, we structurally analyzed Arp4 and Arp8 from Saccharomyces cerevisiae and tested their biochemical effects on actin assembly and disassembly. The solution structures of isolated Arp4 and Arp8 indicate them to be monomeric and the crystal structure of ATP-Arp4 reveals several intriguing differences to actin that explain why Arp4 does not form filaments itself. Remarkably, Arp4, assisted by Arp8, influences actin polymerization in vitro and is able to depolymerize actin filaments by complex formation with monomeric ADP-actin via the barbed end. Our data suggest a likely assembly between actin and Arp4, explaining how nuclear actin is held in a discrete complex within the INO80 chromatin remodeler.
Actin complexes in the cell nucleus: new stones in an old field
Histochemistry and Cell Biology, 2010
Actin is a well-known protein that has shown a myriad of activities in the cytoplasm. However, recent findings of actin involvement in nuclear processes are overwhelming. Actin complexes in the nucleus range from very dynamic chromatin-remodeling complexes to structural elements of the matrix with single partners known as actin-binding proteins (ABPs). This review summarizes the recent findings of actin-containing complexes in the nucleus. Particular attention is given to key processes like chromatin remodeling, transcription, DNA replication, nucleocytoplasmic transport and to actin roles in nuclear architecture. Understanding the mechanisms involving ABPs will definitely lead us to the principles of the regulation of gene expression performed via concerting nuclear and cytoplasmic processes.