activity-dependent-spatially-localized-mirna-maturation-in-neuronal-dendrites

1、Activity-dependent spatially localized miRNA maturation in neuronal dendrites,by Sivakumar Sambandan, Güney Akbalik, Lisa Kochen, Jennifer Rinne, Josefine Kahlstatt, Caspar Glock, Georgi Tushev, Beatriz Alvarez-Cast

2、elao, Alexander Heckel, and Erin M. Schuman,ScienceVolume 355(6325):634-637February 10, 2017,,Zhang Zhenyu2017/09/26,,,Questions,how a miRNA can effectively regulate a particular target mRNA with a relative low copy n

3、umber? whether the maturation of a miRNA could be regulated in space and time in neurons?,Conclusions,1)Pre-miRNA processing occurs in neuronal dendrites and 2)is stimulated by neural activity at individual synapses, 3)

4、resulting in a local reduction in target protein synthesis.,NanoString,,Geiss, G.K., et al., Direct multiplexed measurement of gene expression with color-coded probe pairs. Nat Biotechnol, 2008. 26(3): p. 317-25.,Pre–miR

5、-181a is present in neuronal dendrites.,,,miR-181a is a neuronal miRNA that is highly expressed in the hippocampus,Pre–miR-181a is present in neuronal dendrites.,,,cultured hippocampal neurons,Pre–miR-181a is present in

6、neuronal dendrites.,,,Immunostaining,pre–miR-181a (white),MAP2 (microtubule-associated protein 2; magenta) identifies the dendrites,hippocampal slice,,,http://www.gene-quantification.de/rnai.html,An inducible probe repor

7、ts the maturation of a miRNA.,,,The activity-dependent maturation of a miRNA.,,,elicited action potentials,Intracellular Ca2+ (mediated by NMDA receptors) is important for dicer activity,,The maturation of this miR

8、NA is dicer activity dependent,,,APV (D,L-2-amino-5-phosphonovaleric acid):NMDA receptor antagonist,Glutamate uncaging by two-photo light,,Local stimulation leads to the rapid and spatially restricted maturation of miR-1

9、81a in dendrites,,,Local stimulation leads to the rapid and spatially restricted maturation of miR-181a in spines.,,,Dendritic spine,Dendritic spine,Puromycin - proximity ligation assay （Puro-PLA）,,tom Dieck, S., et

10、 al., Direct visualization of newly synthesized target proteins in situ. Nat Meth, 2015. 12(5): p. 411-414.,Puromycin is an antibiotic that is a protein synthesis inhibitor by inhibiting translation. Causes premature cha

11、in termination during translation taking place in the ribosome.,The local activity-dependent maturation of miR-181a is associated with a decrease in the translation of a target mRNA, CamKIIα.,,,Ca2+-calmodulin protein ki

12、nase α (CamKIIα): a major protein at excitatory synapses and a target of miR-181a,region of mature miR-181a generated,The local activity-dependent maturation of miR-181a is associated with a decrease in the translation o

13、f a target mRNA, CamKIIα.,,,with the miR-181a seed,without the miR-181a seed,Conclusions,1)Pre-miRNA processing occurs in neuronal dendrites and 2)is stimulated by neural activity at individual synapses, 3)resulting in a