Gating-by-tilt of mechanosensitive membrane channels

We propose an alternative mechanism for the gating of biological membrane channels in response to membrane tension that involves a change in the slope of the membrane near the channel. Under biological membrane tensions we show that the energy difference between the closed (tilted) and open (untilted) states can far exceed kBT and is comparable to what is available under simple ilational gating. Recent experiments demonstrate that membrane leaflet asymmetries (spontaneous curvature) can strong effect the gating of some channels. Such a phenomenon would be more easy to explain under gating-by-tilt, given its novel intrinsic sensitivity to such asymmetry.Comment: 10 pages, 2 figure

Hydrophobic gating of mechanosensitive channel of large conductance evidenced by single-subunit resolution

Mechanosensitive (MS) ion channels are membrane proteins that detect and respond to membrane tension in all branches of life. In bacteria, MS channels prevent cells from lysing upon sudden hypoosmotic shock by opening and releasing solutes and water. Despite the importance of MS channels and ongoing efforts to explain their functioning, the molecular mechanism of MS channel gating remains elusive and controversial. Here we report a method that allows single-subunit resolution for manipulating and monitoring “mechanosensitive channel of large conductance” from Escherichia coli. We gradually changed the hydrophobicity of the pore constriction in this homopentameric protein by modifying a critical pore residue one subunit at a time. Our experimental results suggest that both channel opening and closing are initiated by the transmembrane 1 helix of a single subunit and that the participation of each of the five identical subunits in the structural transitions between the closed and open states is asymmetrical. Such a minimal change in the pore environment seems ideal for a fast and energy-efficient response to changes in the membrane tension.

Vibrational Spectra of a Mechanosensitive Channel

We report the simulated vibrational spectra of a mechanosensitive membrane channel in different gating states. Our results show that while linear absorption is insensitive to structural differences, linear dichroism and sum-frequency generation spectroscopies are sensitive to the orientation of the transmembrane helices, which is changing during the opening process. Linear dichroism cannot distinguish an intermediate structure from the closed structure, but sum-frequency generation can. In addition, we find that two-dimensional infrared spectroscopy can be used to distinguish all three investigated gating states of the mechanosensitive membrane channel.

Clonal karyotype evolution involving ring chromosome 1 with myelodysplastic syndrome subtype RAEB-t progressing into acute leukemia

s Karyotypic evolution is a well-known phenomenon in patients with malignant hernatological disorders during disease progression. We describe a 50-year-old male patient who had originally presented with pancytopenia in October 1992. The diagnosis of a myelodysplastic syndrome (MDS) FAB subtype RAEB-t was established in April 1993 by histological bone marrow (BM) examination, and therapy with low-dose cytosine arabinoside was initiated. In a phase of partial hernatological remission, cytogenetic assessment in August 1993 revealed a ring chromosome 1 in 13 of 21 metaphases beside BM cells with normal karyotypes {[}46,XY,r(1)(p35q31)/46,XY]. One month later, the patient progressed to an acute myeloid leukemia (AML), subtype M4 with 40% BM blasts and cytogenetic examination showed clonal evolution by the appearance of additional numerical aberrations in addition to the ring chromosome{[}46,XY,r(1),+8,-21/45,XY,r(1),+8,-21,-22/46, XY]. Intensive chemotherapy and radiotherapy was applied to induce remission in preparation for allogeneic bone marrow transplantation (BMT) from the patient's HLA-compatible son. After BMT, complete remission was clinically, hematologically and cytogenetically (normal male karyotype) confirmed. A complete hematopoietic chimerism was demonstrated. A relapse in January 1997 was successfully treated using donor lymphocyte infusion and donor peripheral blood stem cells (PB-SC) in combination with GM-CSF as immunostimulating agent in April 1997, and the patient's clinical condition remained stable as of January 2005. This is an interesting case of a patient with AML secondary to MDS. With the ring chromosome 1 we also describe a rare cytogenetic abnormality that predicted the poor prognosis of the patient, but the patient could be cured by adoptive immunotherapy and the application of donor's PB-SC. This case confirms the value of cytogenetic analysis in characterizing the malignant clone in hernatological neoplasias, the importance of controlling the quality of an induced remission and of the detection of a progress of the disease. Copyright (c) 2006 S. Karger AG, Basel

Molecular Biomechanics: The Molecular Basis of How Forces Regulate Cellular Function

Recent advances have led to the emergence of molecular biomechanics as an essential element of modern biology. These efforts focus on theoretical and experimental studies of the mechanics of proteins and nucleic acids, and the understanding of the molecular mechanisms of stress transmission, mechanosensing and mechanotransduction in living cells. In particular, single-molecule biomechanics studies of proteins and DNA, and mechanochemical coupling in biomolecular motors have demonstrated the critical importance of molecular mechanics as a new frontier in bioengineering and life sciences. To stimulate a more systematic study of the basic issues in molecular biomechanics, and attract a broader range of researchers to enter this emerging field, here we discuss its significance and relevance, describe the important issues to be addressed and the most critical questions to be answered, summarize both experimental and theoretical/computational challenges, and identify some short-term and long-term goals for the field. The needs to train young researchers in molecular biomechanics with a broader knowledge base, and to bridge and integrate molecular, subcellular and cellular level studies of biomechanics are articulated.National Institutes of Health (U.S.) (grant UO1HL80711-05 to GB)National Institutes of Health (U.S.) (grant R01GM076689-01)National Institutes of Health (U.S.) (grant R01AR033236-26)National Institutes of Health (U.S.) (grant R01GM087677-01A1)National Institutes of Health (U.S.) (grant R01AI44902)National Institutes of Health (U.S.) (grant R01AI38282)National Science Foundation (U.S.) (grant CMMI-0645054)National Science Foundation (U.S.) (grant CBET-0829205)National Science Foundation (U.S.) (grant CAREER-0955291

MscS-like mechanosensitive channels in plants and microbes

The challenge of osmotic stress is something all living organisms must face as a result of environmental dynamics. Over the past three decades, innovative research and cooperation across disciplines have irrefutably established that cells utilize mechanically gated ion channels to release osmolytes and prevent cell lysis during hypoosmotic stress. Early electrophysiological analysis of the inner membrane of Escherichia coli identified the presence of three distinct mechanosensitive activities. The subsequent discoveries of the genes responsible for two of these activities, the mechanosensitive channels of large (MscL) and small (MscS) conductance, led to the identification of two diverse families of mechanosensitive channels. The latter of these two families, the MscS family, consists of members from bacteria, archaea, fungi, and plants. Genetic and electrophysiological analysis of these family members has provided insight into how organisms use mechanosensitive channels for osmotic regulation in response to changing environmental and developmental circumstances. Furthermore, determining the crystal structure of E. coli MscS and several homologues in several conformational states has contributed to our understanding of the gating mechanisms of these channels. Here we summarize our current knowledge of MscS homologues from all three domains of life and address their structure, proposed physiological functions, electrophysiological behaviors, and topological diversity

Single-Molecule Force Spectroscopy: Experiments, Analysis, and Simulations

International audienceThe mechanical properties of cells and of subcellular components are important to obtain a mechanistic molecular understanding of biological processes. The quantification of mechanical resistance of cells and biomolecules using biophysical methods matured thanks to the development of nanotechnologies such as optical and magnetic tweezers, the biomembrane force probe and atomic force microscopy (AFM). The quantitative nature of force spectroscopy measurements has converted AFM into a valuable tool in biophysics. Force spectroscopy allows the determination of the forces required to unfold protein domains and to disrupt individual receptor/ligand bonds. Molecular simulation as a computational microscope allows investigation of similar biological processes with an atomistic detail. In this chapter, we first provide a step-by-step protocol of force spectroscopy including sample preparation, measurement and analysis of force spectroscopy using AFM and its interpretation in terms of available theories. Next, we present the background for molecular dynamics (MD) simulations focusing on steered molecular dynamics (SMD) and the importance of bridging of computational tools with experimental technique