Plasma membrane tension is an important feature that determines the cell shape and influences processes such as cell motility, spreading, endocytosis and exocytosis. Unconventional class 1 myosins are potent regulators of plasma membrane tension because they physically link the plasma membrane with adjacent cytoskeleton. We identified nuclear myosin 1 (NM1) - a putative nuclear isoform of myosin 1c (Myo1c) - as a new player in the field. Although having specific nuclear functions, NM1 localizes predominantly to the plasma membrane. Deletion of NM1 causes more than a 50% increase in the elasticity of the plasma membrane around the actin cytoskeleton as measured by atomic force microscopy. This higher elasticity of NM1 knock-out cells leads to 25% higher resistance to short-term hypotonic environment and rapid cell swelling. In contrast, overexpression of NM1 in wild type cells leads to an additional 30% reduction of their survival. We have shown that NM1 has a direct functional role in the cytoplasm as a dynamic linker between the cell membrane and the underlying cytoskeleton, regulating the degree of effective plasma membrane tension.

• MeSH
• Cell Membrane metabolism   MeSH
• Cell Nucleus metabolism   MeSH
• Exocytosis physiology   MeSH
• Fibroblasts cytology   metabolism   MeSH
• HeLa Cells MeSH
• Cells, Cultured MeSH
• Skin cytology   metabolism   MeSH
• Humans MeSH
• Actin Cytoskeleton metabolism   MeSH
• Myosin Type I metabolism   MeSH
• Mice, Knockout MeSH
• Mice MeSH
• Cell Movement MeSH
• Cell Shape MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Myosin Type I MeSH

Paxillin (PXN) is a focal adhesion protein that has been implicated in signal transduction from the extracellular matrix. Recently, it has been shown to shuttle between the cytoplasm and the nucleus. When inside the nucleus, paxillin promotes cell proliferation. Here, we introduce paxillin as a transcriptional regulator of IGF2 and H19 genes. It does not affect the allelic expression of the two genes; rather, it regulates long-range chromosomal interactions between the IGF2 or H19 promoter and a shared distal enhancer on an active allele. Specifically, paxillin stimulates the interaction between the enhancer and the IGF2 promoter, thus activating IGF2 gene transcription, whereas it restrains the interaction between the enhancer and the H19 promoter, downregulating the H19 gene. We found that paxillin interacts with cohesin and the mediator complex, which have been shown to mediate long-range chromosomal looping. We propose that these interactions occur at the IGF2 and H19 gene cluster and are involved in the formation of loops between the IGF2 and H19 promoters and the enhancer, and thus the expression of the corresponding genes. These observations contribute to a mechanistic explanation of the role of paxillin in proliferation and fetal development.

• Keywords
• Cohesin, Enhancer, H19, IGF2, Imprinting, Paxillin,
• MeSH
• Hep G2 Cells MeSH
• Chromosomal Proteins, Non-Histone genetics   MeSH
• Extracellular Matrix genetics   MeSH
• Focal Adhesions genetics   MeSH
• Genomic Imprinting genetics   MeSH
• Insulin-Like Growth Factor II biosynthesis   genetics   MeSH
• Cohesins MeSH
• Humans MeSH
• DNA Methylation genetics   MeSH
• Paxillin administration & dosage   MeSH
• Cell Proliferation drug effects   genetics   MeSH
• Promoter Regions, Genetic MeSH
• Cell Cycle Proteins genetics   MeSH
• RNA, Long Noncoding biosynthesis   genetics   MeSH
• Signal Transduction drug effects   MeSH
• Fetal Development genetics   MeSH
• Gene Expression Regulation, Developmental MeSH
• Enhancer Elements, Genetic MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Chromosomal Proteins, Non-Histone MeSH
• H19 long non-coding RNA MeSH Browser
• IGF2 protein, human MeSH Browser
• Insulin-Like Growth Factor II MeSH
• Paxillin MeSH
• Cell Cycle Proteins MeSH
• RNA, Long Noncoding MeSH

BACKGROUND: Nuclear myosin I (NM1) is a nuclear isoform of the well-known "cytoplasmic" Myosin 1c protein (Myo1c). Located on the 11(th) chromosome in mice, NM1 results from an alternative start of transcription of the Myo1c gene adding an extra 16 amino acids at the N-terminus. Previous studies revealed its roles in RNA Polymerase I and RNA Polymerase II transcription, chromatin remodeling, and chromosomal movements. Its nuclear localization signal is localized in the middle of the molecule and therefore directs both Myosin 1c isoforms to the nucleus. METHODOLOGY/PRINCIPAL FINDINGS: In order to trace specific functions of the NM1 isoform, we generated mice lacking the NM1 start codon without affecting the cytoplasmic Myo1c protein. Mutant mice were analyzed in a comprehensive phenotypic screen in cooperation with the German Mouse Clinic. Strikingly, no obvious phenotype related to previously described functions has been observed. However, we found minor changes in bone mineral density and the number and size of red blood cells in knock-out mice, which are most probably not related to previously described functions of NM1 in the nucleus. In Myo1c/NM1 depleted U2OS cells, the level of Pol I transcription was restored by overexpression of shRNA-resistant mouse Myo1c. Moreover, we found Myo1c interacting with Pol II. The ratio between Myo1c and NM1 proteins were similar in the nucleus and deletion of NM1 did not cause any compensatory overexpression of Myo1c protein. CONCLUSION/SIGNIFICANCE: We observed that Myo1c can replace NM1 in its nuclear functions. Amount of both proteins is nearly equal and NM1 knock-out does not cause any compensatory overexpression of Myo1c. We therefore suggest that both isoforms can substitute each other in nuclear processes.

• MeSH
• Cell Nucleus metabolism   MeSH
• DNA Primers genetics   MeSH
• Phenotype * MeSH
• Genotype MeSH
• Immunoprecipitation MeSH
• Myosin Type I genetics   metabolism   MeSH
• Mice, Knockout MeSH
• Mice MeSH
• Plasmids genetics   MeSH
• Reverse Transcriptase Polymerase Chain Reaction MeSH
• Protein Isoforms genetics   metabolism   MeSH
• Blotting, Western MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA Primers MeSH
• Myo1c protein, mouse MeSH Browser
• Myosin Type I MeSH
• Protein Isoforms MeSH