Cytoplasmic Dynein ATPase Activity Is Regulated by Dynactin-dependent Phosphorylation

155

128

We have previously uncovered roles for phospholipase D (PLD) and an unknown cytosolic protein in the formation of cytosolic lipid droplets using a cell-free system. In this report, PLD1 has been identified as the relevant isoform, and extracellular signal-regulated kinase 2 (ERK2) as the cytosolic protein. Increased expression of PLD1 increased lipid droplet formation whereas knockdown of PLD1 using siRNA was inhibitory. A role for ERK2 in basal lipid droplet formation was revealed by overexpression or microinjection, and ablation by siRNA knockdown or pharmacological inhibition. Similar manipulations of other Map kinases such as ERK1, JNK1 or JNK2 and p38α or p38β were without effect. Insulin stimulated the formation of lipid droplets and this stimulation was inhibited by knockdown of PLD1 (by siRNA) and by inhibition or knockdown (by siRNA) of ERK2. Inhibition of ERK2 eliminated the effect of PLD1 on lipid droplet formation without affecting PLD1 activity, suggesting that PLD1 functions upstream of ERK2. ERK2 increased the phosphorylation of dynein which increased the amount of the protein on ADRP-containing lipid droplets. Microinjection of antibodies to dynein strongly inhibited the formation of lipid droplets, demonstrating that dynein has a central role in this formation. Thus dynein is a possible target for ERK2.

Section: Discussionmentioning

confidence: 99%

“…It is well known that the activity of dynein is highly dependent on phosphorylation reactions (King, 2000;Kumar et al, 2000). Moreover, differential phosphorylation is one of the mechanisms behind the generation of the isoforms of the cytoplasmic dynein that are essential for the interaction between dynein and its correct cargo (King, 2000).…”

Section: Discussionmentioning

confidence: 99%

PLD1 and ERK2 regulate cytosolic lipid droplet formation

Andersson

Boström

Ericson

et al. 2006

155

128

“…In ssm4∆ cells Dhc1p can no longer accumulate at that point, and therefore might not be able to pull on microtubules. The dynactin complex is also known to regulate dynein motor activity by directly influencing its ATPase activity (Kumar et al, 2000) and increasing its processivity (King and Schroer, 2000). Ssm4p might therefore regulate dynein both by activating its motor activity and by localising it to the microtubule tips, where it can exert its pulling force.…”

Section: Discussionmentioning

confidence: 99%

The p150-Glued Ssm4p regulates microtubular dynamics and nuclear movement in fission yeast

Niccoli

Yamashita

Nurse

et al. 2004

During vegetative growth of the fission yeastSchizosaccharomyces pombe, microtubules nucleate from multiple microtubule organising centres (MTOCs) close to the nucleus, polymerising until they reach the end of the cell and then shrinking back to the cell centre. In response to mating pheromone, S. pombe undergoes a morphological switch from a vegetative to a shmooing growth pattern. The switch in growth mode is paralleled by a switch in microtubular dynamics. Microtubules nucleate mostly from a single MTOC and pull on the ends of the cell to move the nucleus back and forth. This movement continues after cellular and nuclear fusion in the zygote and is important to ensure correct chromosome pairing, recombination and segregation during meiosis. Here we show that Ssm4p, a p150-Glued protein, is induced specifically in response to pheromone and is required for this nuclear movement. Ssm4p is associated with the cytoplasmic dynein complex and together with the CLIP-170 homologue Tip1p regulates dynein heavy chain localisation. We also show that Ssm4p collaborates with Tip1p in establishing the shmooing microtubular array.

“…Dynactin consists of p150 Glued , p50/dynamitin, p24 (also known as DCTN1, DCTN2 and DCTN3 in vertebrates), and a short filament of the actin-related protein Arp1 (also known as ACTR1A in vertebrates) and the actin capping proteins (Schroer, 2004). Dynactin enhances the processivity of dynein movement along microtubules in vitro (Culver-Hanlon et al, 2006;King and Schroer, 2000) and activates dynein (Kumar et al, 2000). Moreover, dynactin also acts as binding sites for cargo associated with the dynein motor.…”

Section: Introductionmentioning

confidence: 99%

Dynactin and Num1 cooperate to establish the cortical anchoring of cytoplasmic dynein inS. pombe

Fujita

Yamashita

Yamamoto

2015

Chromosome movement during meiosis is crucial for homologous pairing and meiotic recombination. During meiotic prophase in fission yeast, rapid nuclear migration is dependent on cytoplasmic dynein, which is anchored to the cell cortex and pulls microtubules, thereby driving nuclear migration. However, the precise mechanisms underlying dynein localization and activation remain unclear. Here, we identified three subunits of dynactin in fission yeast: Arp1, Mug5 and Jnm1 (also known as Mug1). These subunits transiently colocalized with dynein foci at the cell cortex and were essential for the cortical anchoring of dynein. Cortical factor Num1 (also known as Mcp5), which was also required for dynein anchoring, bound to dynein independently of dynactin. Whereas Num1 suppressed the sliding of dynein foci along the cortex, Arp1, Mug5 and Jnm1 were involved in the regulation of shrinkage and bundling of microtubules. From these data, we propose that dynein anchoring is established by cooperation of transient assembly of dynactin and function of Num1 at the cell cortex.