Actin forms the dendritic nucleation network and undergoes rapid polymerization-depolymerization cycles in lamellipodia. To elucidate the mechanism of actin disassembly, we characterized molecular kinetics of the major filament end-binding proteins Arp2/3 complex and capping protein (CP) using single-molecule speckle microscopy. We have determined the dissociation rates of Arp2/3 and CP as 0.048 and 0.58 s−1, respectively, in lamellipodia of live XTC fibroblasts. This CP dissociation rate is three orders of magnitude faster than in vitro. CP dissociates slower from actin stress fibers than from the lamellipodial actin network, suggesting that CP dissociation correlates with actin filament dynamics. We found that jasplakinolide, an actin depolymerization inhibitor, rapidly blocked the fast CP dissociation in cells. Consistently, the coexpression of LIM kinase prolonged CP speckle lifetime in lamellipodia. These results suggest that cofilin-mediated actin disassembly triggers CP dissociation from actin filaments. We predict that filament severing and end-to-end annealing might take place fairly frequently in the dendritic nucleation actin arrays.
mDia1, a Rho effector, belongs to the Formin family of proteins, which shares the conserved tandem FH1-FH2 unit structure. Formins including mDia1 accelerate actin nucleation while interacting with actin filament fast-growing ends. Here our single-molecule imaging revealed fast directional movement of mDia1 FH1-FH2 for tens of microns in living cells. The movement of mDia1 FH1-FH2 was blocked by actin-perturbing drugs, and the speed of mDia1 FH1-FH2 movement appeared to correlate with actin elongation rates. In vitro, mDia1 FH1-FH2 associated persistently with the growing actin barbed end. mDia1 probably moves processively along the growing end of actin filaments in cells, and Formins may be a molecular motility machinery that is independent from motor proteins.
or most of its history, the superconductivity of strontium ruthenate (Sr 2 RuO 4) (ref. 1) has been understood in terms of an odd-parity two-component order parameter with equal-spin pairing in the RuO 2 planes: p x ± ip y (refs. 2-5). This order parameter is chiral: the Cooper pairs have angular momentum l = ±1. The evidence for chirality comes from the zero-field muon spin relaxation (ZF-μSR) data 6 , observation of a non-zero Kerr rotation below the critical temperature T c (ref. 7) and signs in the junction experiments of domains in the superconducting state 8,9 , while evidence for equal-spin pairing came from the absence of a change in the Knight shift below T c in nuclear magnetic resonance 10 and polarized neutron scattering 11 measurements. The Knight shift is related to the spin susceptibility; in conventional opposite-spin-pairing superconductors, it is suppressed below T c. However, in new measurements, it has been found that the Knight shift is, in fact, suppressed below T c (refs. 12-14), by a magnitude that is unlikely to be reconcilable with equal-spin pairing. This revision has called into question a number of other results on Sr 2 RuO 4. It raises a particular challenge for experiments that indicate chirality, because opposite-spin pairing implies an even-parity momentum-space gap structure. If the order parameter is constrained to be even parity, chiral, and composed of components that are degenerate on the tetragonal lattice of Sr 2 RuO 4 , the only possibility is d xz ± id yz order 15. Under conventional understanding, this is a highly unlikely order parameter because it
DIAPH1 encodes human DIA1, a formin protein that elongates unbranched actin. The c.3634+1G>T DIAPH1 mutation causes autosomal dominant nonsyndromic sensorineural hearing loss, DFNA1, characterized by progressive deafness starting in childhood. The mutation occurs near the C‐terminus of the diaphanous autoregulatory domain (DAD) of DIA1, which interacts with its N‐terminal diaphanous inhibitory domain (DID), and may engender constitutive activation of DIA1. However, the underlying pathogenesis that causes DFNA1 is unclear. We describe a novel patient‐derived DIAPH1 mutation (c.3610C>T) in two unrelated families, which results in early termination prior to a basic amino acid motif (RRKR 1204–1207) at the DAD C‐terminus. The mutant DIA1(R1204X) disrupted the autoinhibitory DID‐DAD interaction and was constitutively active. This unscheduled activity caused increased rates of directional actin polymerization movement and induced formation of elongated microvilli. Mice expressing FLAG‐tagged DIA1(R1204X) experienced progressive deafness and hair cell loss at the basal turn and had various morphological abnormalities in stereocilia (short, fused, elongated, sparse). Thus, the basic region of the DAD mediates DIA1 autoinhibition; disruption of the DID‐DAD interaction and consequent activation of DIA1(R1204X) causes DFNA1.
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