Kinases are 1. 7% of all human genes Kinases are 1.7% of all human genes. 95% are serine kinases, threonine and tyrosine are other amino acids that can be phosphorylated. PO4 is negatively charged, changes protein charge which can change its conformation and functional activity.

Figure 23-3

2nd msg cAMP cGMP DAG Ca2+/Cam 1st msg Growth factors Most kinases are maintained in an inactive state, require signal to activate catalytic domains. 4 subunits 2R, 2C.

GSK3b mediator of wnt pathway, wnt signaling allows GSK3b to free up and activate b-catenin which can enter the nucleus and affect gene transcription. Wnt signaling very complex and involved in everything from development and cell fate specification to synapse formation to synaptic plasticity.

Phosphatases just as important as kinases Phosphatases just as important as kinases! Many proteins activated by dephosphorylation and Ppases must end receptor mediated activation to keep signal spatially and temporally relevant.

Jena Bioscience

PPAses are vital: inhibition (okadaic acid) leads to tumor growth PPAses are vital: inhibition (okadaic acid) leads to tumor growth. Inhibition of ppases also helps us understand roles of ppases in the brain

PP1 natural (endogenous) inhibitor DARPP-32 and others PP1 natural (endogenous) inhibitor DARPP-32 and others. All inhibitors are themselves phosphorylated and Po4 of DARPP-32 activates its phosphatase inhibitory activity.

DARPP-32 pretty restricted to D1 receptor expressing neurons (striatum). DA (and other) signaling leads to activation of DARPP-32 activity via PKA (maintains phosphorylation state by inhibiting phosphatase). Glutamate via Ca2+ and CamK inhibits DARPP-32, promoting phosphatase activity.

Different phosphorylation sites reflect different outcomes Different phosphorylation sites reflect different outcomes. Thr 34 phosphorylation by PKA is important for phosphatase inhibition but thr 75 by CamK or CDK5 blocks phosphorylation by PKA at Thr 34, thereby releasing inhibition on PP1 and leading to dephosphorylation of substrates.

Neural properties affected by phosphorylation: presynaptic, postsynaptic and nuclear effects

Neurotransmitter synthesis: PO4 of TH increases its activity—increase affinity for tetrahydrobiopterin cofactor, decrease affinity for end products. Allow modulation of CA synthesis to a variety of external stimuli.

(B) Schematic of the molecular machinery mediating Ca2+-triggered vesicle fusion. The drawing depicts a segment of a docked synaptic vesicle on the top right and the presynaptic active zone in the middle. The three functional elements of the neurotransmitter release machinery are depicted from right to left. On the right, the core fusion machine composed of the SNARE/SM protein complex is shown; this machine comprises the SNARE proteins synaptobrevin/VAMP, syntaxin-1, and SNAP-25 and the SM protein Munc18-1. The Ca2+ sensor synaptotagmin-1 is depicted in the middle; it is composed of a short intravesicular sequence, a single transmembrane region, and two cytoplasmic C2 domains that bind Ca2+, and it functions using complexin (bound to the SNARE complex) as an assistant. The active zone protein complex containing RIM, Munc13, and RIM-BP and a Ca2+ channel in the presynaptic plasma membrane is shown on the left. In this protein complex, RIM binding to specific target proteins coordinates all three functions of the active zone: RIM binding to vesicular rab proteins (Rab3 and Rab27 isoforms) mediates vesicle docking; RIM binding to the central priming factor Munc13 activates vesicle priming; and RIM binding to the Ca2+ channel, both directly and indirectly via RIM-BP, recruits the Ca2+ channels within 100 nm of the docked vesicles for fast excitation-secretion coupling. The overall design of the neurotransmitter release machinery depicted here enables in a single nanodevice fast and efficient triggering of release in response to an action potential by combining a fusion machine with a Ca2+ trigger and an active zone protein complex that positions all elements into appropriate proximity (modified from Kaeser et al., 2011). Phosphorylation critical for synaptic plasticity, LTP. LTP lost in CamK and PKA -/- mice, as well as RIM-1-/- mice. RIM1 is a substrate for PKA Sudhof, 2013

RIM a substrate for PKA

Phosphorylation of synapsin allows vesicles to move into the readily releasable pool, dock and fuse. Recycling or endocytosis is mediated by clatherin and another family of phosphoproteins called dephosphoins—amphiphysin and dynamin above. Dephosphoins are coordinately dephosphorylated by calcineurin phosphatase and must be rephosphorylated to start the next cycle of endocytosis. Substrates for CDK 5 and dynamin also a substrate of PKC.

Dephosphorylation critical for returning vesicles to readily releaseable pool.

Effects on channels by kinases.

Effects on gene transcription by kinases Effects on gene transcription by kinases. General review of transcription.

CREB-cyclic amp response element binding protein a critical convergence point of many kinases.

Synaptic plasticity: PP1 loss of function promotes LTP; is necessary for LTD. High frequency stimulation leads to excess Ca2+ entry and activation of CamKII and AC (CaM sensitive isoforms). cAMP activates PKA, which phosphorylates Inhibitor 1, allowing it to inhibit PP1. Blocking PP1 allows CaMKII to achieve Ca2+ independent activation through autoactivation, and activate targets such as NOS and AMPA receptors.