Transcription profiling by high throughput sequencing of Arabidopsis not2 mutant seedlings

MicroRNAs (miRNAs) play key regulatory roles in numerous developmental and physiological processes in animals and plants. The elaborate mechanism of miRNA biogenesis involves transcription and multiple processing steps. Here, we report the identification of a pair of evolutionarily conserved NOT2_3_5 domain containing proteins, NOT2a and NOT2b (previously known as At-NOT2 and VIP2, respectively), as components involved in Arabidopsis miRNA biogenesis. NOT2 was identified by its interaction with the Piwi/Ago/Zwille (PAZ) domain of Dicer-like 1 (DCL1), an interaction that is conserved between rice and Arabidopsis. Inactivation of both NOT2 genes in Arabidopsis caused severe defects in male gametophytes, and weak lines show pleiotropic defects reminiscent of miRNA pathway mutants. Impairment of NOT2s decreases the accumulation of pri-miRNAs and mature miRNAs and increases DCL1-containing nuclear speckle number in vivo. In addition, NOT2b protein interacts with Pol II and other miRNA processing factors including two cap-binding proteins, CBP80/ABH1, CBP20 and SERRATE (SE). Therefore, these results suggest that NOT2 proteins promote MIR transcription and facilitate efficient DCL1 recruitment in Arabidopsis. Examination of transcriptome of not2 and wild type by RNA-seq.