Kazuko Fujitani

Mutations in the Novel Membrane Protein Spinster Interfere with Programmed Cell Death and Cause Neural Degeneration in Drosophila melanogaster

ABSTRACT Mutations in the spin gene are characterized by an extraordinarily strong rejection behavior of female flies in response to male courtship. They are also accompanied by decreases in the viability, adult life span, and oviposition rate of the flies. Inspin mutants, some oocytes and adult neural cells undergo degeneration, which is preceded by reductions in programmed cell death of nurse cells in ovaries and of neurons in the pupal nervous system, respectively. The central nervous system (CNS) of spinmutant flies accumulates autofluorescent lipopigments with characteristics similar to those of lipofuscin. The spinlocus generates at least five different transcripts, with only two of these being able to rescue the spin behavioral phenotype; each encodes a protein with multiple membrane-spanning domains that are expressed in both the surface glial cells in the CNS and the follicle cells in the ovaries. Orthologs of the spin gene have also been identified in a number of species from nematodes to humans. Analysis of the spin mutant will give us new insights into neurodegenerative diseases and aging.

N‐acetyltransferase ARD1‐NAT1 regulates neuronal dendritic development

ARD1 and NAT1 constitute an N‐acetyltransferase complex where ARD1 holds the enzymatic activity of the complex. The ARD1–NAT1 complex mediates N‐terminal acetylation of nascent polypeptides that emerge from ribosomes after translation. ARD1 may also acetylate the internal lysine residues of proteins. Although ARD1 and NAT1 have been found in the brain, the physiological role and substrates of the ARD1–NAT1 complex in neurons remain unclear. Here we investigated role of N‐acetyltransferase activity in the process of neuronal development. Expression of ARD1 and NAT1 increased during dendritic development, and both proteins colocalized with microtubules in dendrites. The ARD1–NAT1 complex displayed acetyltransferase activity against a purified microtubule fraction in vitro. Inhibition of the complex limited the dendritic extension of cultured neurons. These findings suggest that the ARD1–NAT1 complex has acetyltransferase activity against microtubules in dendrites. Regulation by acetyltransferase activity is a novel mechanism that is required for dendritic arborization during neuronal development.

The microtubule destabilizer stathmin mediates the development of dendritic arbors in neuronal cells

The regulation of microtubule dynamics is important for the appropriate arborization of neuronal dendrites during development, which in turn is critical for the formation of functional neural networks. Here we show that stathmin, a microtubule destabilizing factor, is downregulated at both the expression and activity levels during cerebellar development, and this down-regulation contributes to dendritic arborization. Stathmin overexpression drastically limited the dendritic growth of cultured Purkinje cells. The stathmin activity was suppressed by neural activity and CaMKII-dependent phosphorylation at Ser16, which led to dendritic arborization. Stathmin phosphorylation at Ser16 was mediated by the activation of voltage-gated calcium channels and metabotropic glutamate receptor 1. Although overexpression of SCG10, a member of the stathmin family, also limited the dendritic arborization, SCG10 did not mediate the CaMKII regulation of dendritic development. These results suggest that calcium elevation activates CaMKII, which in turn phosphorylates stathmin at Ser16 to stabilize dendritic microtubules. siRNA knockdown of endogenous stathmin significantly reduced dendritic growth in Purkinje cells. Thus, these data suggest that proper regulation of stathmin activity is a key factor for controlling the dendritic microtubule dynamics that are important for neuronal development.

Phorbol esters promote postsynaptic accumulation of Vesl-1S/Homer-1a protein: Phorbol esters promote postsynaptic accumulation of Vesl-1S

We examined effects of phorbol esters on the amount and the subcellular distribution of the activity‐regulated protein Vesl‐1S/Homer‐1a in cultured hippocampal neurons. Major Vesl‐1S immunoreactivity (IR) was detected throughout neuronal somata under control conditions. Bath application of phorbol esters, PMA and PDBu resulted in the increase in the amount of Vesl‐1S proteins and promoted punctate distribution of Vesl‐1S IR at the cortical regions of the neuronal somata. Immunofluorescent observations using antisynaptophysin and anti‐Vesl‐1S antibodies, and electron microscopic observations, revealed that Vesl‐1S accumulated at postsynaptic regions following PMA application. Membrane depolarization with high concentrations of external potassium also promoted the punctate distribution of Vesl‐1S IR. These results demonstrate that phorbol‐triggered reaction cascades result in the accumulation of Vesl‐1S protein at postsynaptic regions, and suggest that these phorbol effects may mimic those caused by synaptic activities.