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ECHO-liveFISH: in vivo RNA labeling reveals dynamic regulation of nuclear RNA foci in living tissues

2015; Oxford University Press; Volume: 43; Issue: 19 Linguagem: Inglês

10.1093/nar/gkv614

1362-4962

Ikumi Oomoto, Asuka Suzuki‐Hirano, Hiroki Umeshima, Yong-Woon Han, Hiroyuki Yanagisawa, Peter M. Carlton, Yoshie Harada, Mineko Kengaku, Akimitsu Okamoto, Tomomi Shimogori, Dan Ohtan Wang,

RNA Interference and Gene Delivery

Elucidating the dynamic organization of nuclear RNA foci is important for understanding and manipulating these functional sites of gene expression in both physiological and pathological states. However, such studies have been difficult to establish in vivo as a result of the absence of suitable RNA imaging methods. Here, we describe a high-resolution fluorescence RNA imaging method, ECHO-liveFISH, to label endogenous nuclear RNA in living mice and chicks. Upon in vivo electroporation, exciton-controlled sequence-specific oligonucleotide probes revealed focally concentrated endogenous 28S rRNA and U3 snoRNA at nucleoli and poly(A) RNA at nuclear speckles. Time-lapse imaging reveals steady-state stability of these RNA foci and dynamic dissipation of 28S rRNA concentrations upon polymerase I inhibition in native brain tissue. Confirming the validity of this technique in a physiological context, the in vivo RNA labeling did not interfere with the function of target RNA nor cause noticeable cytotoxicity or perturbation of cellular behavior.