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The nucleolar protein SAHY1 is involved in pre-rRNA processing and normal plant growth.

ABSTRACT: Although the nucleolus is involved in ribosome biogenesis, the functions of numerous nucleolus-localized proteins remain unclear. In this study, we genetically isolated Arabidopsis thaliana salt hypersensitive mutant 1 (sahy1), which exhibits slow growth, short roots, pointed leaves, and sterility. SAHY1 encodes an uncharacterized protein that is predominantly expressed in root tips, early developing seeds, and mature pollen grains and is mainly restricted to the nucleolus. Dysfunction of SAHY1 primarily causes the accumulation of 32S, 18S-A3, and 27SB pre-rRNA intermediates. Coimmunoprecipitation experiments further revealed the interaction of SAHY1 with ribosome proteins and ribosome biogenesis factors. Moreover, sahy1 mutants are less sensitive to protein translation inhibitors and show altered expression of structural constituents of ribosomal genes and ribosome subunit profiles, reflecting the involvement of SAHY1 in ribosome composition and ribosome biogenesis. Analyses of ploidy, S-phase cell cycle progression, and auxin transport and signaling indicated the impairment of mitotic activity, translation of auxin transport carrier proteins, and expression of the auxin-responsive marker DR5::GFP in the root tips or embryos of sahy1 plants. Collectively, these data demonstrate that SAHY1, a nucleolar protein involved in ribosome biogenesis, plays critical roles in normal plant growth in association with auxin transport and signaling.

SUBMITTER: Hsu PJ

PROVIDER: S-EPMC8133687 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Although the nucleolus is known to be involved in the plant stress response, the underlying mechanisms remain unclear. In the present study, we genetically isolated and characterized the Arabidopsis thaliana salt hypersensitive mutant 1 (sahy1), which exhibits slow growth, short roots, and sterility. SAHY1 encodes an unknown protein that is predominantly expressed in root tips and mature pollens. Subcellular localization further demonstrated that SAHY1 is a nucleolar protein. Ploidy analysis and detection of S-phase cell cycle progression indicated that mitosis is reduced in the root tips of sahy1 plants. Furthermore, levels of reactive oxygen species (ROS) are reduced in the mutant. In addition to exhibiting changes in venation pattern, palisade mesophyll cells, the auxin response, and fertility, the sahy1 mutant is resistant to the protein translation inhibitors, streptomycin and cycloheximide, and has altered expression profiles of ribosome-related genes, reflecting the involvement of SAHY1 in ribosome assembly. Using a combination of transcriptome and gene coexpression clustering analyses, we identified a striking functional module in which SPIRAL 1-LIKE 4 (SPL4), which encodes a microtubule-localized protein, mediates the salt response that acts by modulating dynamic microtubule instability. Supporting evidence indicates that sahy1 is resistant to a microtubule-destabilizing drug (oryzalin), that the cortical microtubule array in sahy1 exhibits changes in dynamic instability, and that the spr1/spl2/spl3/spl4 quadruple mutant is oversensitive to salt. Collectively, these data demonstrate that a novel nucleolar protein, SAHY1, plays critical roles in salt sensitivity, plant growth, and development by integrating the diverse functions of its downstream genes. As the sahy1 mutant plants showed slow growth, short roots, and sterility, the aim of this experiment is to examine gene expression profile in root tissues mediated by SAHY1. The data obtained from this study was compared with chip database derived from root tissues and performed an analysis of gene coexpression clustering.

Project description:The biogenesis of ribosomes requires tightly controlled transcription and processing of pre-rRNA which comprises ribosomal RNAs forming the core of large and small ribosomal subunits. Early steps of the pre-rRNA processing and assembly of the ribosomal subunits require a large set of proteins that perform folding and nucleolytic cleavage of pre-rRNAs in the nucleoli. Structure and functions of proteins involved in the pre-rRNA processing have been extensively studied in the budding yeast S. cerevisiae. Functional characterization of their human homologues is complicated by the complexity of mammalian ribosomes and increased number of protein factors involved in the ribosomal biogenesis. Homologues of human nucleolar protein SURF6 from yeast and mouse, Rrp14 and Surf6, respectively, had been shown to be involved in the early steps of pre-rRNA processing. Rrp14 works as RNA chaperone in complex with proteins Ssf1 and Rrp15. Human SURF6 knockdown and overexpression were used to clarify a role of SURF6 in the early steps of pre-rRNA processing in human cell lines HeLa and HTC116. By analyzing the abundance of the rRNA precursors in cells with decreased level or overexpression of SURF6, we demonstrated that human SURF6 is involved in the maturation of rRNAs from both small and large ribosomal subunits. Changes in the SURF6 level caused by knockdown or overexpression of the protein do not result in the death of HeLa cells in contrast to murine embryonic fibroblasts, but significantly alter the distribution of cells among the phases of the cell cycle. SURF6 knockdown in both p53 sufficient and p53 deficient HCT116 human cancer cells results in elongation of G0/G1 and shortening of G2/M phase. This surprising result suggests p53 independence of SURF6 effects on the cell cycle and possible multiple functions of SURF6. Our data point to the shift from pathway 1 to pathway 2 of the rRNA biogenesis caused by the SURF6 knockdown and its likely association with p53 pathway.

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The nucleolar protein SAHY1 is involved in pre-rRNA processing and normal plant growth.

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