While the dynamin GTPase Drp1 plays a critical role during mitochondrial fission, mechanisms controlling its recruitment to fission sites are unclear. A current assumption is that cytosolic Drp1 is recruited directly to fission sites immediately prior to fission. Using live-cell microscopy, we find evidence for a different model, progressive maturation of Drp1 oligomers on mitochondria through incorporation of smaller mitochondrially-bound Drp1 units. Maturation of a stable Drp1 oligomer does not forcibly lead to fission. Drp1 oligomers also translocate directionally along mitochondria. Ionomycin, a calcium ionophore, causes rapid mitochondrial accumulation of actin filaments followed by Drp1 accumulation at the fission site, and increases fission rate. Inhibiting actin polymerization, myosin IIA, or the formin INF2 reduces both un-stimulated and ionomycin-induced Drp1 accumulation and mitochondrial fission. Actin filaments bind purified Drp1 and increase GTPase activity in a manner that is synergistic with the mitochondrial protein Mff, suggesting a role for direct Drp1/actin interaction. We propose that Drp1 is in dynamic equilibrium on mitochondria in a fission-independent manner, and that fission factors such as actin filaments target productive oligomerization to fission sites.DOI: http://dx.doi.org/10.7554/eLife.11553.001

Quantitative and reproducible data can be obtained from surface-based DNA sensors if variations in the conformation and surface density of immobilized single-stranded DNA capture probes are minimized. Both the conformation and surface density can be independently and deterministically controlled by taking advantage of the preferential adsorption of adenine nucleotides (dA) on gold, as previously demonstrated using a model system in Opdahl, A.; Petrovykh, D. Y.; Kimura-Suda, H.; Tarlov, M. J.; Whitman, L. J. Proc. Natl. Acad. Sci. U.S.A. 2007, 104, 9-14. Here, we describe the immobilization and subsequent hybridization properties of a 15-nucleotide DNA probe sequence that has additional m adenine nucleotides, (dA)(m), at the 5' end. Quantitative analysis of immobilization and hybridization for these probes indicates that the (dA)(m) block preferentially adsorbs on gold, forcing the probe portion of the strand to adopt an upright conformation suited for efficient hybridization. In addition, a wide range of probe-to-probe lateral spacing can be achieved by coimmobilizing the probe DNA with a lateral spacer, a strand of k adenine nucleotides, (dA)(k). Altering either the length or relative concentration of the (dA)(k) spacers added during probe immobilization controls the average surface density of probes; the density of probes, in turn, systematically modulates their hybridization with solution targets.

Mitochondrial dynamics, including fusion, fission and translocation, are crucial to cellular homeostasis, with roles in cellular polarity, stress response and apoptosis. Mitochondrial fission has received particular attention, owing to links with several neurodegenerative diseases. A central player in fission is the cytoplasmic dynaminrelated GTPase Drp1, which oligomerizes at the fission site and hydrolyzes GTP to drive membrane ingression. Drp1 recruitment to the outer mitochondrial membrane (OMM) is a key regulatory event, which appears to require a pre-constriction step in which the endoplasmic reticulum (ER) and mitochondrion interact extensively, a process termed ERMD (ER-associated mitochondrial division). It is unclear how ER-mitochondrial contact generates the force required for pre-constriction or why pre-constriction leads to Drp1 recruitment. Recent results, however, show that ERMD might be an actin-based process in mammals that requires the ER-associated formin INF2 upstream of Drp1, and that myosin II and other actinbinding proteins might be involved. In this Commentary, we present a mechanistic model for mitochondrial fission in which actin and myosin contribute in two ways; firstly, by supplying the force for preconstriction and secondly, by serving as a coincidence detector for Drp1 binding. In addition, we discuss the possibility that multiple fission mechanisms exist in mammals.

A tendency in cell biology is to divide and conquer. For example, decades of painstaking work have led to an understanding of endoplasmic reticulum (ER) and Golgi structure, dynamics, and transport. In parallel, cytoskeletal researchers have revealed a fantastic diversity of structure and cellular function in both actin and microtubules. Increasingly, these areas overlap, necessitating an understanding of both organelle and cytoskeletal biology. This review addressesconnections between the actin/microtubule cytoskeletons and organelles in animal cells, focusing on threetopics: ER structure/function, ER-to-Golgi transport; and Golgi structure/function. Making these connections has been challenging, due to 1) the small sizes and dynamic characteristics of some components, 2) the fact that organelle-specific cytoskeleton can easily be obscured by more abundant cytoskeletal structures, and 3) the difficulties in imaging membranes and cytoskeleton simultaneously, especially at the ultra-structural level. One major concept is that the cytoskeleton is frequently used to generate force for membrane movement, with two potential consequences: translocation of the organelle, or deformation of the organelle membrane. While initially discussing issues common to metazoan cells in general, we subsequently highlight specific features of neurons, since these highly polarized cells present unique challenges for organellar distribution and dynamics.

Actin stimulates oligomerization and mitochondrial accumulation of Drp1, a mitochondrial fission protein. Drp1 binds actin filaments in an unusually dynamic manner that is strongly influenced by guanine nucleotide.