Publications by authors named "Jaakko Lehtimäki"

11 Publications

  • Page 1 of 1

Generation of stress fibers through myosin-driven reorganization of the actin cortex.

Elife 2021 Jan 28;10. Epub 2021 Jan 28.

HiLIFE Institute of Biotechnology, University of Helsinki, Helsinki, Finland.

Contractile actomyosin bundles, stress fibers, govern key cellular processes including migration, adhesion, and mechanosensing. Stress fibers are thus critical for developmental morphogenesis. The most prominent actomyosin bundles, ventral stress fibers, are generated through coalescence of pre-existing stress fiber precursors. However, whether stress fibers can assemble through other mechanisms has remained elusive. We report that stress fibers can also form without requirement of pre-existing actomyosin bundles. These structures, which we named cortical stress fibers, are embedded in the cell cortex and assemble preferentially underneath the nucleus. In this process, non-muscle myosin II pulses orchestrate the reorganization of cortical actin meshwork into regular bundles, which promote reinforcement of nascent focal adhesions, and subsequent stabilization of the cortical stress fibers. These results identify a new mechanism by which stress fibers can be generated from the actin cortex and establish role for stochastic myosin pulses in the assembly of functional actomyosin bundles.
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http://dx.doi.org/10.7554/eLife.60710DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7877910PMC
January 2021

Assembly of Peripheral Actomyosin Bundles in Epithelial Cells Is Dependent on the CaMKK2/AMPK Pathway.

Cell Rep 2020 03;30(12):4266-4280.e4

Section of Pathology, Department of Veterinary Biosciences, University of Helsinki, Helsinki, Finland. Electronic address:

Defects in the maintenance of intercellular junctions are associated with loss of epithelial barrier function and consequent pathological conditions, including invasive cancers. Epithelial integrity is dependent on actomyosin bundles at adherens junctions, but the origin of these junctional bundles is incompletely understood. Here we show that peripheral actomyosin bundles can be generated from a specific actin stress fiber subtype, transverse arcs, through their lateral fusion at cell-cell contacts. Importantly, we find that assembly and maintenance of peripheral actomyosin bundles are dependent on the mechanosensitive CaMKK2/AMPK signaling pathway and that inhibition of this route leads to disruption of tension-maintaining actomyosin bundles and re-growth of stress fiber precursors. This results in redistribution of cellular forces, defects in monolayer integrity, and loss of epithelial identity. These data provide evidence that the mechanosensitive CaMKK2/AMPK pathway is critical for the maintenance of peripheral actomyosin bundles and thus dictates cell-cell junctions through cellular force distribution.
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http://dx.doi.org/10.1016/j.celrep.2020.02.096DOI Listing
March 2020

UNC-45a promotes myosin folding and stress fiber assembly.

J Cell Biol 2017 12 20;216(12):4053-4072. Epub 2017 Oct 20.

Institute of Biotechnology, University of Helsinki, Helsinki, Finland

Contractile actomyosin bundles, stress fibers, are crucial for adhesion, morphogenesis, and mechanosensing in nonmuscle cells. However, the mechanisms by which nonmuscle myosin II (NM-II) is recruited to those structures and assembled into functional bipolar filaments have remained elusive. We report that UNC-45a is a dynamic component of actin stress fibers and functions as a myosin chaperone in vivo. UNC-45a knockout cells display severe defects in stress fiber assembly and consequent abnormalities in cell morphogenesis, polarity, and migration. Experiments combining structured-illumination microscopy, gradient centrifugation, and proteasome inhibition approaches revealed that a large fraction of NM-II and myosin-1c molecules fail to fold in the absence of UNC-45a. The remaining properly folded NM-II molecules display defects in forming functional bipolar filaments. The C-terminal UNC-45/Cro1/She4p domain of UNC-45a is critical for NM-II folding, whereas the N-terminal tetratricopeptide repeat domain contributes to the assembly of functional stress fibers. Thus, UNC-45a promotes generation of contractile actomyosin bundles through synchronized NM-II folding and filament-assembly activities.
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http://dx.doi.org/10.1083/jcb.201703107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5716280PMC
December 2017

Lipid Bilayer-Gated Mesoporous Silica Nanocarriers for Tumor-Targeted Delivery of Zoledronic Acid in Vivo.

Mol Pharm 2017 09 10;14(9):3218-3227. Epub 2017 Aug 10.

Pharmaceutical Sciences Laboratory, Faculty of Science and Engineering, Åbo Akademi University , Turku 20520, Finland.

Zoledronic acid (ZOL) is a nitrogen-containing bisphosphonate used for the treatment of bone diseases and calcium metabolism. Anticancer activity of ZOL has been established, but its extraskeletal effects are limited due to its rapid uptake and accumulation to bone hydroxyapatite. In this work, we report on the development of tethered lipid bilayer-gated mesoporous silica nanocarriers (MSNs) for the incorporation, retention, and intracellular delivery of ZOL. The in vitro anticancer activity of ZOL-loaded nanocarriers was evaluated by cell viability assay and live-cell imaging. For in vivo delivery, the nanocarriers were tagged with folic acid to boost the affinity for breast cancer cells. Histological examination of the liver revealed no adverse off-target effects stemming from the nanocarriers. Importantly, nonspecific accumulation of ZOL within bone was not observed, which indicated in vivo stability of the tethered lipid bilayers. Further, the intravenously administered ZOL-loaded nanocarriers showed tumor growth suppression in breast cancer xenograft-bearing mice.
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http://dx.doi.org/10.1021/acs.molpharmaceut.7b00519DOI Listing
September 2017

Toll-like receptor 9 expression is associated with breast cancer sensitivity to the growth inhibitory effects of bisphosphonates in vitro and in vivo.

Oncotarget 2016 Dec;7(52):87373-87389

Department of Chemistry, University of Alabama at Birmingham, Birmingham, AL, U.S.A.

Bisphosphonates are standard treatments for bone metastases. When given in the adjuvant setting, they reduce breast cancer mortality and recurrence in bone but only among post-menopausal patients. Optimal drug use would require biomarker-based patient selection. Such biomarkers are not yet in clinical use. Based on the similarities in inflammatory responses to bisphosphonates and Toll-like receptor (TLR) agonists, we hypothesized that TLR9 expression may affect bisphosphonate responses in cells. We compared bisphosphonate effects in breast cancer cell lines with low or high TLR9 expression. We discovered that cells with decreased TLR9 expression are significantly more sensitive to the growth-inhibitory effects of bisphosphonates in vitro and in vivo. Furthermore, cancer growth-promoting effects seen with some bisphosphonates in some control shRNA cells were not detected in TLR9 shRNA cells. These differences were not associated with inhibition of Rap1A prenylation or p38 phosphorylation, which are known markers for bisphosphonate activity. However, TLR9 shRNA cells exhibited increased sensitivity to ApppI, a metabolite that accumulates in cells after bisphosphonate treatment. We conclude that decreased TLR9-expression sensitizes breast cancer cells to the growth inhibitory effects of bisphosphonates. Our results suggest that TLR9 should be studied as a potential biomarker for adjuvant bisphosphonate sensitivity among breast cancer patients.
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http://dx.doi.org/10.18632/oncotarget.13570DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5349995PMC
December 2016

Actin Filament Structures in Migrating Cells.

Handb Exp Pharmacol 2017 ;235:123-152

Institute of Biotechnology, University of Helsinki, 56, 00014, Helsinki, Finland.

Cell migration is necessary for several developmental processes in multicellular organisms. Furthermore, many physiological processes such as wound healing and immunological events in adult animals are dependent on cell migration. Consequently, defects in cell migration are linked to various diseases including immunological disorders as well as cancer progression and metastasis formation. Cell migration is driven by specific protrusive and contractile actin filament structures, but the types and relative contributions of these actin filament arrays vary depending on the cell type and the environment of the cell. In this chapter, we introduce the most important actin filament structures that contribute to mesenchymal and amoeboid cell migration modes and discuss the mechanisms by which the assembly and turnover of these structures are controlled by various actin-binding proteins.
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http://dx.doi.org/10.1007/164_2016_28DOI Listing
September 2017

Spheroid culture of LuCaP 136 patient-derived xenograft enables versatile preclinical models of prostate cancer.

Clin Exp Metastasis 2016 Apr 12;33(4):325-37. Epub 2016 Feb 12.

Department of Urology, Stanford University School of Medicine, Stanford, CA, 94305, USA.

LuCaP serially transplantable patient-derived xenografts (PDXs) are valuable preclinical models of locally advanced or metastatic prostate cancer. Using spheroid culture methodology, we recently established cell lines from several LuCaP PDXs. Here, we characterized in depth the features of xenografts derived from LuCaP 136 spheroid cultures and found faithful retention of the phenotype of the original PDX. In vitro culture enabled luciferase transfection into LuCaP 136 spheroids, facilitating in vivo imaging. We showed that LuCaP 136 spheroids formed intratibial, orthotopic, and subcutaneous tumors when re-introduced into mice. Intratibial tumors responded to castration and were highly osteosclerotic. LuCaP 136 is a realistic in vitro-in vivo preclinical model of a subtype of bone metastatic prostate cancer.
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http://dx.doi.org/10.1007/s10585-016-9781-2DOI Listing
April 2016

Bidirectional Interplay between Vimentin Intermediate Filaments and Contractile Actin Stress Fibers.

Cell Rep 2015 Jun 28;11(10):1511-8. Epub 2015 May 28.

Institute of Biotechnology, P.O. Box 56, 00014 University of Helsinki, Helsinki, Finland. Electronic address:

The actin cytoskeleton and cytoplasmic intermediate filaments contribute to cell migration and morphogenesis, but the interplay between these two central cytoskeletal elements has remained elusive. Here, we find that specific actin stress fiber structures, transverse arcs, interact with vimentin intermediate filaments and promote their retrograde flow. Consequently, myosin-II-containing arcs are important for perinuclear localization of the vimentin network in cells. The vimentin network reciprocally restricts retrograde movement of arcs and hence controls the width of flat lamellum at the leading edge of the cell. Depletion of plectin recapitulates the vimentin organization phenotype of arc-deficient cells without affecting the integrity of vimentin filaments or stress fibers, demonstrating that this cytoskeletal cross-linker is required for productive interactions between vimentin and arcs. Collectively, our results reveal that plectin-mediated interplay between contractile actomyosin arcs and vimentin intermediate filaments controls the localization and dynamics of these two cytoskeletal systems and is consequently important for cell morphogenesis.
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http://dx.doi.org/10.1016/j.celrep.2015.05.008DOI Listing
June 2015

Human monoclonal Fab and human plasma antibodies to carbamyl-epitopes cross-react with malondialdehyde-adducts.

Immunology 2014 Mar;141(3):416-30

Department of Medical Microbiology and Immunology, Institute of Diagnostics, University of Oulu, Oulu, Finland; Medical Research Center Oulu, Oulu, Finland; NordLab Oulu, Oulu University Hospital, Oulu, Finland.

Oxidized low-density lipoprotein (OxLDL) plays a crucial role in the development of atherosclerosis. Carbamylated LDL has been suggested to promote atherogenesis in patients with chronic kidney disease. Here we observed that plasma IgG and IgM antibodies to carbamylated epitopes were associated with IgG and IgM antibodies to oxidation-specific epitopes (ρ = 0·65-0·86, P < 0·001) in healthy adults, suggesting a cross-reaction between antibodies recognizing carbamyl-epitopes and malondialdehyde (MDA)/malondialdehyde acetaldehyde (MAA) -adducts. We used a phage display technique to clone a human Fab antibody that bound to carbamylated LDL and other carbamylated proteins. Anti-carbamyl-Fab (Fab106) cross-reacted with oxidation-specific epitopes, especially with MDA-LDL and MAA-LDL. We showed that Fab106 bound to apoptotic Jurkat cells known to contain these oxidation-specific epitopes, and the binding was competed with soluble carbamylated and MDA-/MAA-modified LDL and BSA. In addition, Fab106 was able to block the uptake of carbamyl-LDL and MDA-LDL by macrophages and stained mouse atherosclerotic lesions. The observed cross-reaction between carbamylated and MDA-/MAA-modified LDL and its contribution to enhanced atherogenesis in uraemic patients require further investigation.
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http://dx.doi.org/10.1111/imm.12204DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3930379PMC
March 2014

Natural antibodies of newborns recognize oxidative stress-related malondialdehyde acetaldehyde adducts on apoptotic cells and atherosclerotic plaques.

Int Immunol 2013 Oct 30;25(10):575-87. Epub 2013 Jul 30.

Department of Medical Microbiology and Immunology, Institute of Diagnostics, Faculty of Medicine, University of Oulu, 90014 Oulu, Finland.

Malondialdehyde acetaldehyde (MAA) adducts are generated under oxidative stress and shown to be highly immunogenic. Our aim was to investigate the recognition of MAA adducts by human natural antibodies in newborns before or at the time of full-term pregnancy. Plasma samples of pre-term (n = 11) and full-term (n = 36) newborns were enriched in specific IgM binding to MAA adducts compared with the maternal plasma IgM levels. Umbilical cord blood lymphocyte phage display library was generated to clone Fabs that specifically recognized MAA adducts without cross-reactivity to malondialdehyde. Fab clones from the antibody libraries of the pre-term and full-term newborns showed high sequence homology to the germline genes encoding the variable regions of antibodies, confirming that these Fabs represented the natural antibody repertoire of human fetuses. The MAA-specific umbilical cord blood Fabs bound to apoptotic human endothelial cells and the binding was efficiently competed with MAA adducts. The MAA-specific Fabs also recognized epitopes on advanced atherosclerotic lesions, and the uptake of infrared (IR)-labeled MAA-low-density lipoprotein by mouse J774A.1 macrophages was significantly reduced in the presence of these Fabs. In conclusion, MAA adducts were identified as one of the major antigenic targets for human natural antibodies already before the time of birth. MAA-specific natural antibodies are suggested to regulate apoptotic cell clearance starting from fetal development and to participate in the immunomodulation of atherosclerosis development during adulthood.
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http://dx.doi.org/10.1093/intimm/dxt022DOI Listing
October 2013

Carbamyl adducts on low-density lipoprotein induce IgG response in LDLR-/- mice and bind plasma autoantibodies in humans under enhanced carbamylation.

Antioxid Redox Signal 2013 Oct 1;19(10):1047-62. Epub 2013 Mar 1.

1 Department of Medical Microbiology & Immunology, Institute of Diagnostics, University of Oulu , Oulu, Finland .

Aims: Post-translational modification of proteins via carbamylation predicts increased risk for coronary artery disease. Uremia and smoke exposure are known to increase carbamylation. The aim was to investigate the role of carbamylated low-density lipoprotein (LDL) immunization on antibody formation and atherogenesis in LDL receptor-deficient (LDLR-/-) mice, and to study autoantibodies to carbamylated proteins in humans with carbamylative load.

Results: LDLR-/- mice immunized with carbamylated mouse LDL (msLDL; n=10) without adjuvant showed specific immunoglobulin G (IgG) antibody levels to carbamyl-LDL and malondialdehyde-modified LDL (MDA-LDL) but not to oxidized LDL or native LDL. Immunization did not influence the atherosclerotic plaque area compared with control LDLR-/- mice immunized with native msLDL (n=10) or phosphate-buffered saline (n=11). Humans with high plasma urea levels, as well as smokers, had increased IgG autoantibody levels to carbamyl-modified proteins compared to the subjects with normal plasma urea levels, or to nonsmokers.

Innovation: Carbamyl-LDL induced specific IgG antibody response cross-reactive with MDA-LDL in mice. IgG antibodies to carbamyl-LDL were also found in human plasma and related to conditions known to have increased carbamylation, such as uremia and smoking. Plasma antibodies to carbamylated proteins may serve as new indicator of in vivo carbamylation.

Conclusion: These data give insight into mechanisms of in vivo humoral recognition of post-translationally modified structures. Humoral IgG immune response to carbamylated proteins is suggested to play a role in conditions leading to enhanced carbamylation, such as uremia and smoking.
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http://dx.doi.org/10.1089/ars.2012.4535DOI Listing
October 2013