Keratins Stabilize Hemidesmosomes through Regulation of β4-Integrin Turnover

Epidermal integrity and wound healing depend on remodeling of cell-matrix contacts including hemidesmosomes. Mutations in β4-integrin and plectin lead to severe epidermolysis bullosa (EB). Whether mutations in keratins K5 or K14, which cause EB simplex, also compromise cell-matrix adhesion through altering hemidesmosomal components is not well investigated. In particular, the dependence of β4-integrin endocytosis and turnover on keratins remains incompletely understood. Here, we show that the absence of keratins causes loss of plectin-β4-integrin interaction and elevated β4-integrin phosphorylation at Ser1354 and Ser1362. This triggered a caveolin-dependent endocytosis of β4-integrin but not of other integrins through Rab5 and Rab11 compartments in keratinocytes. Expressing a phospho–deficient β4-integrin mutant reduces β4-integrin endocytosis and rescues plectin localization in keratin–free cells. β4-integrin phosphorylation in the absence of keratins resulted from elevated Erk1/2 activity downstream of increased EGFR and PKCα signaling. Further, increased Erk1/2 phosphorylation and altered plectin localization occur in keratin–deficient mouse epidermis in vivo. Strikingly, expression of the K14-R125P EBS mutant also resulted in plectin mislocalization and elevated β4-integrin turnover, suggesting disease relevance. Our data underscore a major role of keratins in controlling β4-integrin endocytosis involving a plectin-Erk1/2-dependent mechanism relevant for epidermal differentiation and pathogenesis

Tunable singlet-triplet splitting in a few-electron Si/SiGe quantum dot

We measure the excited-state spectrum of a Si/SiGe quantum dot as a function of in-plane magnetic field, and we identify the spin of the lowest three eigenstates in an effective two-electron regime. The singlet-triplet splitting is an essential parameter describing spin qubits, and we extract this splitting from the data. We find it to be tunable by lateral displacement of the dot, which is realized by changing two gate voltages on opposite sides of the device. We present calculations showing the data are consistent with a spectrum in which the first excited state of the dot is a valley-orbit state.Comment: 4 pages with 3 figure

Vimentin intermediate filaments control actin stress fiber assembly through GEF-H1 and RhoA

The actin and intermediate filament cytoskeletons contribute to numerous cellular processes, including morphogenesis, cytokinesis and migration. These two cytoskeletal systems associate with each other, but the underlying mechanisms of this interaction are incompletely understood. Here, we show that inactivation of vimentin leads to increased actin stress fiber assembly and contractility, and consequent elevation of myosin light chain phosphorylation and stabilization of tropomyosin-4.2 (see Geeves et al., 2015). The vimentin-knockout phenotypes can be rescued by re-expression of wild-type vimentin, but not by the non-filamentous ` unit length form' vimentin, demonstrating that intact vimentin intermediate filaments are required to facilitate the effects on the actin cytoskeleton. Finally, we provide evidence that the effects of vimentin on stress fibers are mediated by activation of RhoA through its guanine nucleotide exchange factor GEF-H1 (also known as ARHGEF2). Vimentin depletion induces phosphorylation of the microtubule-associated GEF-H1 on Ser886, and thereby promotes RhoA activity and actin stress fiber assembly. Taken together, these data reveal a new mechanism by which intermediate filaments regulate contractile actomyosin bundles, and may explain why elevated vimentin expression levels correlate with increased migration and invasion of cancer cells.Peer reviewe

Transport through an impurity tunnel coupled to a Si/SiGe quantum dot

Achieving controllable coupling of dopants in silicon is crucial for operating donor-based qubit devices, but it is difficult because of the small size of donor-bound electron wavefunctions. Here we report the characterization of a quantum dot coupled to a localized electronic state, and we present evidence of controllable coupling between the quantum dot and the localized state. A set of measurements of transport through this device enable the determination of the most likely location of the localized state, consistent with an electronically active impurity in the quantum well near the edge of the quantum dot. The experiments we report are consistent with a gate-voltage controllable tunnel coupling, which is an important building block for hybrid donor and gate-defined quantum dot devices.Comment: 5 pages, 3 figure

Two-axis control of a singlet-triplet qubit with an integrated micromagnet

The qubit is the fundamental building block of a quantum computer. We fabricate a qubit in a silicon double quantum dot with an integrated micromagnet in which the qubit basis states are the singlet state and the spin-zero triplet state of two electrons. Because of the micro magnet, the magnetic field difference $\Delta B$ between the two sides of the double dot is large enough to enable the achievement of coherent rotation of the qubit's Bloch vector about two different axes of the Bloch sphere. By measuring the decay of the quantum oscillations, the inhomogeneous spin coherence time $T_{2}^{*}$ is determined. By measuring $T_{2}^{*}$ at many different values of the exchange coupling $J$ and at two different values of $\Delta B$, we provide evidence that the micromagnet does not limit decoherence, with the dominant limits on $T_{2}^{*}$ arising from charge noise and from coupling to nuclear spins.Comment: 10 pages, 9 figure

Quantum control and process tomography of a semiconductor quantum dot hybrid qubit

The similarities between gated quantum dots and the transistors in modern microelectronics - in fabrication methods, physical structure, and voltage scales for manipulation - have led to great interest in the development of quantum bits (qubits) in semiconductor quantum dots. While quantum dot spin qubits have demonstrated long coherence times, their manipulation is often slower than desired for important future applications, such as factoring. Further, scalability and manufacturability are enhanced when qubits are as simple as possible. Previous work has increased the speed of spin qubit rotations by making use of integrated micromagnets, dynamic pumping of nuclear spins, or the addition of a third quantum dot. Here we demonstrate a new qubit that offers both simplicity - it requires no special preparation and lives in a double quantum dot with no added complexity - and is very fast: we demonstrate full control on the Bloch sphere with $\pi$-rotation times less than 100 ps in two orthogonal directions. We report full process tomography, extracting high fidelities equal to or greater than 85% for X-rotations and 94% for Z-rotations. We discuss a path forward to fidelities better than the threshold for quantum error correction.Comment: 6 pages, excluding Appendi

Cytoskeletal vimentin regulates cell size and autophagy through mTORC1 signaling

The nutrient-activated mTORC1 (mechanistic target of rapamycin kinase complex 1) signaling pathway determines cell size by controlling mRNA translation, ribosome biogenesis, protein synthesis, and autophagy. Here, we show that vimentin, a cytoskeletal intermediate filament protein that we have known to be important for wound healing and cancer progression, determines cell size through mTORC1 signaling, an effect that is also manifested at the organism level in mice. This vimentin-mediated regulation is manifested at all levels of mTOR downstream target activation and protein synthesis. We found that vimentin maintains normal cell size by supporting mTORC1 translocation and activation by regulating the activity of amino acid sensing Rag GTPase. We also show that vimentin inhibits the autophagic flux in the absence of growth factors and/or critical nutrients, demonstrating growth factor-independent inhibition of autophagy at the level of mTORC1. Our findings establish that vimentin couples cell size and autophagy through modulating Rag GTPase activity of the mTORC1 signaling pathway

Vimentin takes a hike – Emerging roles of extracellular vimentin in cancer and wound healing

Vimentin is a cytoskeletal protein important for many cellular processes, including proliferation, migration, invasion, stress resistance, signaling, and many more. The vimentin-deficient mouse has revealed many of these functions as it has numerous severe phenotypes, many of which are found only following a suitable challenge or stress. While these functions are usually related to vimentin as a major intracellular protein, vimentin is also emerging as an extracellular protein, exposed at the cell surface in an oligomeric form or secreted to the extracellular environment in soluble and vesicle-bound forms. Thus, this review explores the roles of the extracellular pool of vimentin (eVIM), identified in both normal and pathological states. It focuses specifically on the recent advances regarding the role of eVIM in wound healing and cancer. Finally, it discusses new technologies and future perspectives for the clinical application of eVIM

Evaluation of aldehyde dehydrogenase 1 promoter polymorphisms identified in human populations

BACKGROUND: Cytosolic aldehyde dehydrogenase, or ALDH1A1, functions in ethanol detoxification, metabolism of neurotransmitters, and synthesis of retinoic acid. Because the promoter region of a gene can influence gene expression, the ALDH1A1 promoter regions were studied to identify polymorphism, to assess their functional significance, and to determine whether they were associated with a risk for developing alcoholism. METHODS: Sequence analysis was performed in the promoter region by using Asian, Caucasian, and African American subjects. The resulting polymorphisms were assessed for frequency in Asian, Caucasian, Jewish, and African American populations and tested for associations with alcohol dependence in Asian and African American populations of alcoholics and controls. The functional significance of each polymorphism was determined through in vitro expression analysis by using HeLa and HepG2 cells. RESULTS: Two polymorphisms, a 17 base pair (bp) deletion (-416/-432) and a 3 bp insertion (-524), were discovered in the ALDH1A1 promoter region: ALDH1A1*2 and ALDH1A1*3, respectively. ALDH1A1*2 was observed at frequencies of 0.035, 0.023, 0.023, and 0.012 in the Asian, Caucasian, Jewish, and African American populations, respectively. ALDH1A1*3 was observed only in the African American population, at a frequency of 0.029. By using HeLa and HepG2 cells for in vitro expression, the activity of the luciferase reporter gene was significantly decreased after transient transfection of ALDH1A1*3-luciferase compared with the wild-type construct ALDH1A1*1-luciferase. In an African American population, a trend for higher frequencies of the ALDH1A1*2 and ALDH1A1*3 alleles was observed in a population of alcoholics (p = 0.03 and f = 0.12, respectively) compared with the control population. CONCLUSIONS: ALDH1A1*2 and ALDH1A1*3 may influence ALDH1A1 gene expression. Both ALDH1A1*2 and ALDH1A1*3 produce a trend in an African American population that may be indicative of an association with alcoholism; however, more samples are required to validate this observation. The underlying mechanisms contributing to these trends are still unknown