I-Phosphorylation yenza kusebenze isilawuli sokukhula esiyinhloko i-DELLA ku-Arabidopsis ngokuphromotha ukuhlotshaniswa kwe-histone H2A ne-chromatin.

Amaprotheni e-DELLA ayingcweti elondolozwayoizilawuli zokukhulaezidlala indima ebalulekile ekulawuleni ukuthuthukiswa kwezitshalo ekuphenduleni izimpawu zangaphakathi nezemvelo. I-DELLA isebenza njengesilawuli sokulotshiweyo futhi iqashelwa ukukhomba abathuthukisi ngokubophezela kuzinto zokuloba (TFs) kanye ne-histone H2A ngesizinda sayo se-GRAS. Ucwaningo lwakamuva lubonise ukuthi ukuzinza kwe-DELLA kulawulwa ngemva kokuhumusha ngokusebenzisa izindlela ezimbili: i-polyubiquitination eyenziwa yi-phytohormone gibberellin, okuholela ekuwohlokeni kwayo okusheshayo, kanye nokuhlanganiswa kwe-ubiquitin-like modifiers (SUMO) ukuze kwandiswe ukuqoqwa kwayo. Ngaphezu kwalokho, umsebenzi we-DELLA ulawulwa ngamandla ngama-glycosylations amabili ahlukene: ukusebenzisana kwe-DELLA-TF kuthuthukiswa yi-O-fucosylation kodwa kuvinjwe ukuguqulwa kwe-O-linked N-acetylglucosamine (O-GlcNAc). Kodwa-ke, indima ye-DELLA phosphorylation isalokhu ingacacile, njengoba izifundo zangaphambilini zibonise imiphumela ephikisanayo, kusukela kulabo ababonisa ukuthi i-phosphorylation ikhuthaza noma inciphisa ukuchithwa kwe-DELLA kwabanye okubonisa ukuthi i-phosphorylation ayithinti ukuzinza kwayo. Lapha, sikhomba amasayithi e-phosphorylation ku-REPRESSORiga1-3(I-RGA, i-AtDELLA) ihlanzwe ku-Arabidopsis thaliana ngokuhlaziywa kwe-mass spectrometry futhi ikhombisa ukuthi i-phosphorylation yama-peptide amabili e-RGA ezifundeni ze-PolyS kanye ne-PolyS/T ikhuthaza umsebenzi obophayo we-H2A futhi othuthukisiwe we-RGA. Ukuhlangana kwe-RGA nabaphromotha abaqondiwe. Ngokuphawulekayo, i-phosphorylation ayithinti ukusebenzisana kwe-RGA-TF noma ukuzinza kwe-RGA. Ucwaningo lwethu lwembula indlela yamangqamuzana lapho i-phosphorylation idala umsebenzi we-DELLA.
Ukucacisa indima ye-phosphorylation ekulawuleni umsebenzi we-DELLA, kubalulekile ukukhomba izindawo ze-DELLA phosphorylation ku-vivo futhi wenze ukuhlaziya okusebenzayo ezitshalweni. Ngokucwenga okuhlobene kokukhishwe kwezitshalo okulandelwa ukuhlaziya kwe-MS/MS, sihlonze ama-phosphosite ambalwa ku-RGA. Ngaphansi kwezimo zokuntuleka kwe-GA, i-RHA phosphorylation iyanda, kodwa i-phosphorylation ayithinti ukuzinza kwayo. Okubalulekile, ukuhlolwa kwe-co-IP ne-ChIP-qPCR kwembula ukuthi i-phosphorylation endaweni ye-PolyS/T ye-RGA ikhuthaza ukusebenzisana kwayo ne-H2A kanye nokuhlangana kwayo nabagqugquzeli abaqondisiwe, okuveza indlela i-phosphorylation eyenza ngayo umsebenzi we-RGA.
I-RGA iqashelwa ukukhomba i-chromatin ngokusebenzisana kwesizinda esingaphansi kwe-LHR1 ne-TF bese ibophezela ku-H2A ngesifunda sayo se-PolyS/T kanye nesizinda esingaphansi kwe-PFYRE, okwenza inkimbinkimbi ye-H2A-RGA-TF ukuze kuzinziswe i-RGA. I-Phosphorylation ye-Pep 2 esifundeni se-PolyS/T phakathi kwesizinda se-DELLA nesizinda se-GRAS nge-kinase engaziwa ithuthukisa ukubophezela kwe-RGA-H2A. Iphrotheni eguquguqukayo ye-rgam2A iqeda i-phosphorylation ye-RGA futhi yamukela ukwakheka okuhlukile kwamaprotheni ukuze iphazamise ukubophezela kwe-H2A. Lokhu kubangela ukungasebenzi kahle kokusebenzelana kwe-TF-rgam2A kwesikhashana kanye nokuhlukaniswa kwe-rgam2A ku-chromatin eqondiwe. Lesi sibalo sibonisa kuphela ukucindezela kokulotshiweyo okuphakathi kwe-RGA. Iphethini efanayo ingachazwa ngokwenza kusebenze okulotshiweyo okuphakathi kwe-RGA, ngaphandle kokuthi inkimbinkimbi ye-H2A-RGA-TF izophromotha ukubhalwa kofuzo okuqondiwe futhi i-dephosphorylation ye-rgam2A izokwehlisa ukuloba. Umfanekiso ulungiswe kusuka ku-Huang et al.21.
Yonke idatha yobuningi yahlaziywa ngokwezibalo kusetshenziswa i-Excel, futhi umehluko omkhulu wanqunywa kusetshenziswa ukuhlolwa kuka-t koMfundi. Azikho izindlela zezibalo ezisetshenzisiwe ukuze kutholwe usayizi wesampula kuqala. Ayikho idatha ekhishiwe ekuhlaziyweni; ukuhlolwa akuzange kwenziwe ngokungahleliwe; abacwaningi bebengaphuphutheki ekusabalaliseni idatha ngesikhathi sokuhlolwa nokuhlolwa kwemiphumela. Usayizi wesampula ukhonjiswe kunganekwane yomfanekiso kanye nefayela ledatha yomthombo.
Ukuze uthole ulwazi olwengeziwe mayelana nomklamo wocwaningo, bheka I-Natural Portfolio Report Abstract ehlobene nalesi sihloko.
Idatha ye-mass spectrometry proteomics inikelwe kunhlangano ye-ProteomeXchange ngekhosombe likazakwethu we-PRIDE66 elinesikhombi sedathasethi i-PXD046004. Yonke eminye imininingwane etholwe phakathi nalolu cwaningo yethulwa kokuthi Ulwazi Olwengeziwe, Amafayela Edatha Eyengeziwe, kanye Namafayela Edatha Eluhlaza. Idatha yomthombo inikezwe lesi sihloko.