Numb proteins specify asymmetric cell fates via an endocytosis- and proteasome-independent pathway.

Numb proteins are evolutionarily conserved signaling molecules that make the daughter cells different after asymmetric divisions by segregating to only one daughter. They contain distinct binding motifs for alpha-adaptin (alpha-Ada) and proteins with Eps15 homology (EH) domains, which regulate endocytosis, and for E3 ubiquitin ligases, which target proteins for proteasome-mediated degradation. In Drosophila melanogaster, Numb acts by inhibiting Notch activity to cause a bias in Notch-mediated cell-cell communication. These findings have led to the hypothesis that Numb modulates Notch signaling by using endocytosis and proteasomes to directly reduce Notch protein levels at the cell surface. Here we show that two Drosophila EH proteins, Eps15 homologue 1 (EH1) and the dynamin-associated 160-kDa protein (Dap160), negatively regulate Notch signaling. However, neither elimination of the binding motifs for endocytic proteins nor simultaneous reduction of proteasome activity affects the activity of Numb proteins. Our findings indicate that an endocytosis- and proteasome-independent pathway may mediate Numb signaling in asymmetric cell fate specification.

The mammalian Golgi regulates numb signaling in asymmetric cell division by releasing ACBD3 during mitosis.

Mammalian neural progenitor cells divide asymmetrically to self-renew and produce a neuron by segregating cytosolic Numb proteins primarily to one daughter cell. Numb signaling specifies progenitor over neuronal fates but, paradoxically, also promotes neuronal differentiation. Here we report that ACBD3 is a Numb partner in cell-fate specification. ACBD3 and Numb proteins interact through an essential Numb domain, and the respective loss- and gain-of-function mutant mice share phenotypic similarities. Interestingly, ACBD3 associates with the Golgi apparatus in neurons and interphase progenitor cells but becomes cytosolic after Golgi fragmentation during mitosis, when Numb activity is needed to distinguish the two daughter cells. Accordingly, cytosolic ACBD3 can act synergistically with Numb to specify cell fates, and its continuing presence during the progenitor cell cycle inhibits neuron production. We propose that Golgi fragmentation and reconstitution during cell cycle differentially regulate Numb signaling through changes in ACBD3 subcellular distribution and represent a mechanism for coupling cell-fate specification and cell-cycle progression.

Acetylcholinesterase inhibitors activate septohippocampal GABAergic neurons via muscarinic but not nicotinic receptors.

Acetylcholinesterase (AChE) inhibitors, which increase synaptic levels of available acetylcholine (ACh) by preventing its degradation, are the most extensively prescribed drugs for the treatment of Alzheimer's disease. In animals, AChE inhibitors improve learning and memory, reverse scopolamine-induced amnesia, and produce hippocampal theta rhythm. The medial septum/diagonal band of Broca (MSDB), which maintains hippocampal theta rhythm and associated mnemonic functions via the septohippocampal pathway, is considered a critical locus for mediating the effects of AChE inhibitors. Using electrophysiological recordings and fluorescent labeling techniques to identify living septohippocampal neurons in rat brain slices, we report that AChE inhibitors, in the absence of exogenous ACh, produce a profound excitation in 94% of septohippocampal GABAergic neurons and an inhibition in 24% of septohippocampal cholinergic neurons. The inhibitory and excitatory effects of AChE inhibitors, presumably, occur due to accumulation of ACh that is released locally within the MSDB via axon collaterals of septohippocampal cholinergic neurons. The excitatory effects of AChE inhibitors on septohippocampal GABAergic neurons were blocked by muscarinic but not nicotinic receptor antagonists, especially by the M3 receptor antagonist, 4-diphenylacetoxy-N-methylpiperidine mustard, and not by M1 or M2/M4 muscarinic receptor antagonists. M3 muscarinic receptor mRNA colocalized with the calcium-binding protein, parvalbumin, a marker of septohippocampal GABAergic neurons. These findings may be useful in designing therapeutic strategies that do not depend on endogenous ACh and may therefore be effective in situations where AChE inhibitors cease to be effective, such as in progressive neurodegeneration.