ФоÑоÑелÑÑа ÑаÑÑений â ÑÑо немембÑаннÑе оÑÐ³Ð°Ð½ÐµÐ»Ð»Ñ (ÐÐÐ), обÑазование коÑоÑÑÑ Ð¸Ð½Ð¸ÑииÑÑеÑÑÑ Ð±ÐµÐ»Ð¾Ðº-белковÑми взаимодейÑÑвиÑми Ð¼ÐµÐ¶Ð´Ñ Ð°ÐºÑив-ной ÑоÑмой ÑиÑÐ¾Ñ Ñома B (phyB) и ÑиÑÐ¾Ñ Ñом-взаимодейÑÑвÑÑÑими ÑакÑоÑами (PIF). ÐÑи оÑÐ³Ð°Ð½ÐµÐ»Ð»Ñ Ð¿ÑинимаÑÑ ÑÑаÑÑие в ÑоÑомоÑÑогенезе ÑаÑÑений. Рданной ÑабоÑе бÑл ÑазÑабоÑан Ñ Ð¸Ð¼ÐµÑнÑй белок PIF6-TagYFP-Fus, ÑÑÑановлена его ÑÐºÐ·Ð¾Ð³ÐµÐ½Ð½Ð°Ñ ÑкÑпÑеÑÑÐ¸Ñ Ð² клеÑÐºÐ°Ñ Ð¼Ð»ÐµÐºÐ¾Ð¿Ð¸ÑаÑÑÐ¸Ñ Ð²Ð¼ÐµÑÑе Ñ Ð´ÑÑгим Ñ Ð¸Ð¼ÐµÑнÑм белком phyB-mCherry-Fus, и Ñ Ð¿Ð¾Ð¼Ð¾ÑÑÑ ÐºÐ¾Ð½ÑокалÑной ÑлÑоÑеÑÑенÑной микÑоÑкопии изÑÑена Ð¸Ñ Ð³ÐµÑеÑодимеÑизаÑÐ¸Ñ Ð¸ обÑазование на Ð¸Ñ Ð¾Ñнове ÐÐÐ. РезÑлÑÑаÑÑ Ð¿Ð¾ÐºÐ°Ð·Ð°Ð»Ð¸, ÑÑо облÑÑение ÑаÑÑеÑннÑм ÑвеÑом Ñ Ð´Ð»Ð¸Ð½Ð¾Ð¹ Ð²Ð¾Ð»Ð½Ñ 630 нм индÑÑиÑовало обÑазование и поÑледÑÑÑее ÑвелиÑение ÑазмеÑов ÐÐÐ. СбоÑка и ÑазбоÑка ÑоÑоиндÑÑиÑÑемÑÑ ÐÐРв ÑиÑоплазме клеÑок млекопиÑаÑÑÐ¸Ñ Ð¿Ð¾Ð´ÑинÑÑÑÑÑ Ð·Ð°ÐºÐ¾Ð½Ð°Ð¼, пÑиÑÑÑим конÑенÑÑаÑионно-завиÑÐ¸Ð¼Ð¾Ð¼Ñ ÑÐ°Ð·Ð¾Ð²Ð¾Ð¼Ñ ÑÐ°Ð·Ð´ÐµÐ»ÐµÐ½Ð¸Ñ Ð±Ð¸Ð¾Ð¿Ð¾Ð»Ð¸Ð¼ÐµÑов. РазмеÑÑ ÐÐÐ, обÑазованнÑÑ Ð¸Ð· phyB и PIF6 в клеÑÐºÐ°Ñ Ð¼Ð»ÐµÐºÐ¾Ð¿Ð¸ÑаÑÑÐ¸Ñ , ÑооÑвеÑÑÑвовали ÑазмеÑам Ñак назÑваемÑÑ Â«ÑÐ°Ð½Ð½Ð¸Ñ Â» ÑоÑоÑÐµÐ»ÐµÑ Ð² клеÑÐºÐ°Ñ ÑаÑÑений. ÐÑи ÑезÑлÑÑаÑÑ Ð¿Ð¾Ð·Ð²Ð¾Ð»ÑÑÑ Ð¿ÑедположиÑÑ, ÑÑо пеÑÐ²Ð°Ñ ÑÑÐ°Ð´Ð¸Ñ Ð¾Ð±ÑÐ°Ð·Ð¾Ð²Ð°Ð½Ð¸Ñ ÑоÑоÑÐµÐ»ÐµÑ Ñ ÑаÑÑений Ð¼Ð¾Ð¶ÐµÑ Ð±ÑÑÑ Ð¾Ñнована на ÑвеÑозавиÑимом Ñазовом пеÑÐµÑ Ð¾Ð´Ðµ жидкоÑÑÑ-жидкоÑÑÑ Ð´Ð»Ñ PIF и дÑÑÐ³Ð¸Ñ Ð±ÐµÐ»ÐºÐ¾Ð², коÑоÑÑе могÑÑ ÑпеÑиÑиÑеÑки взаимодейÑÑвоваÑÑ Ñ Ð°ÐºÑивной ÑоÑмой phyB. ÐолÑÑеннÑе Ñ Ð¸Ð¼ÐµÑнÑе белки в пÑинÑипе могÑÑ Ð¸ÑполÑзоваÑÑÑÑ Ð´Ð»Ñ ÑпÑÐ°Ð²Ð»ÐµÐ½Ð¸Ñ ÑбоÑкой и ÑазбоÑкой ÑоÑоиндÑÑиÑÑемÑÑ ÐÐO и, Ñаким обÑазом, Ð´Ð»Ñ ÑегÑлиÑÐ¾Ð²Ð°Ð½Ð¸Ñ ÑазлиÑнÑÑ Ð²Ð½ÑÑÑиклеÑоÑнÑÑ Ð¿ÑоÑеÑÑов в клеÑÐºÐ°Ñ Ð¼Ð»ÐµÐºÐ¾Ð¿Ð¸ÑаÑÑÐ¸Ñ . Plant photobodies are the membrane-less organelles (MLOs) that can be generated by protein-protein interactions between active form of phytochrome B (phyB) and phytochrome-interacting factors (PIFs). These organelles regulate plant photo-morphogenesis. In this work, the chimeric protein PIF6-TagYFP-Fus was developed, its and another developed chimeric proteins phyB-mCherry-Fus exogenous expression in mammalian cells was established, and their heterodimerization and the formation of NMOs based on them were studied using confocal fluorescence microscopy. Results showed that irradiation with diffused 630-nm light induced formation and subsequent increase in sizes of the MLOs. The assembly and disassembly of the photo-inducible MLOs in the mammalian cell cytoplasm obeyed the laws inherent in the concentration-dependent phase separation of biopolymers. The sizes of MLOs formed from phyB and PIF6 in mammalian cells corresponded to the sizes of the so-called âearlyâ photobodies in plant cells. These results suggested that the first step for the formation of plant photobodies might be based on the light-dependent liquid-liquid phase separation of PIFs and other proteins that can specifically interact with the active form of phyB. The developed chimeric proteins in principle can be used to control the assembly and disassembly of photo-inducible MLOs, and thereby to regulate various intracellular processes in mammalian cells.
