Extended Data Fig. 5: Cal-ID/cpTurboID biotinylation and neuronal activity manipulation.
From: Molecular recording of calcium signals via calcium-dependent proximity labeling
(a) cpTurboID biotinylation levels are not associated with neuronal activity. Primary hippocampal neurons at DIV 21. Independent measurements: n = 2. Silenced: 50 µM APV (NMDAR antagonist), 10 µM NBQX (AMPAR antagonist). APV + NBQX: treated overnight. Stimulated: 8 mM CaCl2 was added along with biotin. Labeling: 100 µM biotin was added and labeled for 1 hour. P-S6 is neuronal activity marker. Scale bars: 50 µm. (b) Chemical LTP (cLTP) increases Cal-ID biotinylation. At DIV 21, mouse primary cortical neurons were transferred to aCSF (artificial cerebrospinal fluid) with 500 nM TTX, 20 µM bicuculline, and 1 µM strychnine, equilibrated for 20 minutes. For cLTP condition, Mg2+ was removed and 200 µM glycine was added. 30 minutes incubation with 100 µM biotin for labeling. Independent measurements: n = 2. Numbers on the left: molecular weight (kDa). *: endogenous biotin carrier proteins, #: self-biotinylation of Cal-ID. Numbers at the bottom: densitometry results, * indicates control condition.
