Cell dedifferentiation and organogenesis in vitro require more snRNA than does seedling development in Arabidopsis thaliana

Abstract

Small nuclear RNA (snRNA) is a class of non-coding RNAs that processes pre-mRNA and rRNA. Transcription of abundant snRNA species is regulated by the snRNA activating protein complex (SNAPc), which is conserved among multicellular organisms including plants. SRD2, a putative subunit of SNAPc in Arabidopsis thaliana, is essential for development, and the point mutation srd2-1 causes severe defects in hypocotyl dedifferentiation and de novo meristem formation. Based on phenotypic analysis of srd2-1 mutant plants, we previously proposed that snRNA content is a limiting factor in dedifferentiation in plant cells. Here, we performed functional complementation analysis of srd2-1 using transgenic srd2-1 Arabidopsis plants harboring SRD2 homologs from Populus trichocarpa (poplar), Nicotiana tabacum (tobacco), Oryza sativa (rice), the moss Physcomitrella patens, and Homo sapiens (human) under the control of the Arabidopsis SRD2 promoter. Only rice SRD2 suppressed the faulty tissue culture responses of srd2-1, and restore the snRNA levels; however, interestingly, all SRD2 homologs except poplar SRD2 rescued the srd2-1 defects in seedling development. These findings demonstrated that cell dedifferentiation and organogenesis induced during tissue culture require higher snRNA levels than does seedling development.

Silencing Arabidopsis CARBOXYL‐TERMINAL DOMAIN PHOSPHATASE‐LIKE 4 induces cytokinin‐oversensitive de novo shoot organogenesis

“…In the SRD2‐dependent pathway, both CPL4 RNAi and srd2‐1 compromised snRNA biogenesis (Ohtani and Sugiyama, ; Fukudome et al ., ). However, because the effect of CPL4 RNAi by itself on the total snRNA level was negligible under the standard conditions (Fukudome et al ., ), the inhibitory effects might become significant only in limited processes, such as the formation of the auxin gradient at the developing LRP (Ohtani et al ., ; Ohtani, ). When combined with the srd2‐1 mutation, however, the CPL4 RNAi srd2‐1 calli were no longer able to proliferate at the restrictive temperature (Figure S4), suggesting that the level of functional snRNA pool might have dropped below the threshold to sustain the cell proliferation (Ohtani et al ., ).…”

“…CKH2/PICKLE encodes a SWItch/Sucrose Non‐Fermentable (SWI/SNF) chromatin remodelling factor, which promotes histone H3 K27 trimethylation mark (Zhang et al ., ), and functions synergistically with CKH1 (Furuta et al ., ). SHOOT REDIFFERENTIATION DEFECTIVE 2 ( SRD2 ) is an essential gene for in vitro shoot regeneration from root explants (Yasutani et al ., ), and encodes a homologue of the SNAP50 subunit in the snRNA activator protein complex (SNAPc), suggesting that the proper production of snRNA is essential for dedifferentiation and regeneration of tissues (Ohtani and Sugiyama, ; Ohtani et al ., ). Similarly, a screening for defects in adventitious root formation from shoot explants identified genes that were involved in pre‐rRNA processing ( ROOT INITIATION DEFECTIVE , RID2 and RID3 ) and spliceosome activity ( RID1 , a DEAH‐box helicase homologous to yeast Prp22) (Sugiyama, ; Ohtani et al ., ).…”

Plant snRNP Biogenesis: A Perspective from the Nucleolus and Cajal Bodies

“…Mutations of LSM4 and LSM5, which encode essential core proteins of Sm-like class snRNPs ( Figure 2 ; Achsel et al, 1999 ), and of Tgs1, which hypermethylates the 5′ cap of snRNAs ( Figure 2 ; Mouaikel et al, 2002 ), were reported to enhance plant sensitivities to abiotic stresses, such as salt, drought, and cold stress ( Xiong et al, 2001 ; Zhang et al, 2011 ; Gao et al, 2017 ). Moreover, SHOOT REDIFFERENTIATION DEFECTIVE 2 (SRD2), a subunit of the snRNA-specific transcription activator complex, SNAPc ( Figure 2 ), is required for in vitro dedifferentiation and organogenesis ( Ohtani and Sugiyama, 2005 ; Ohtani et al, 2015 ; Ohtani, 2015 , 2017 ). The disorders of the corresponding mutants can be explained by the misregulation—due to altered RNA processing—of specific genes involved in key processes, i.e., circadian rhythms, stress responses, and auxin polar transport ( Xiong et al, 2001 ; Deng et al, 2010 ; Hong et al, 2010 ; Ohtani et al, 2010 ; Sanchez et al, 2010 ; Zhang et al, 2011 ; Schlaen et al, 2015 ).…”

Alternative splicing of DSP1 enhances snRNA accumulation by promoting transcription termination and recycle of the processing complex

“…Important structural and functional roles of snRNAs in the spliceosome suggest that they may be present in cells at constant amounts (20)(21)(22). However, snRNA levels vary among different organs and developmental stages (2,23,24), and the spatiotemporal regulation of snRNA accumulation is critical for various biological processes (25)(26)(27). For example, the snRNA contents are correlated with the amount of the transcription activator SHOOT REDIFFERENTIATION DEFECTIVE 2 (SRD2), which is strongly expressed in highly proliferated tissues, and impaired function of SRD2 has a stronger impact on the development of these tissues (24).…”