Asymmetric evolution of duplicate genes encoding the CCAAT-binding factor NF-Y in plant genomes

NF-Y is a ubiquitous CCAAT-binding factor composed of NF-YA, NF-YB and NF-YC. Multiple genes encoding NF-Y subunits have been identified in plant genomes. It remains unclear whether the duplicate genes underwent different evolutionary patterns. Likelihood-ratio tests were used to examine whether the amino acid substitution rates are the same between duplicate genes. The influences of selection on evolution were evaluated by comparing the conservative and radical amino acid substitution rates, as well as maximum-likelihood analysis. Some NF-YB and NF-YC duplicates showed significant evidence of asymmetric evolution but not the NF-YA duplicates. Most amino acid replacements in the NF-YB and NF-YC duplicates result in changes in hydropathy, polar requirement and polarity. The physicochemical changes in the sequences of NF-YB seem to be coupled to asymmetric divergence in gene function. Plant NF-Y genes have evolved in different patterns. Relaxed selective constraints following gene duplication are most likely responsible for the unequal evolutionary rates and distinct divergence patterns of duplicate NF-Y genes. Positive selection may have promoted amino acid hydropathy changes in the NF-YC duplicates.     

Recombinant AtNF-YA, a subunit of theArabidopsis CCAAT-binding transcription factor, forms a protein-DNA complex with mammalian NF-YB and NF-YC

The evolutionary conserved CCAAT binding protein NF-Y is a common regulatory DNA binding protein consisting of three distinct subunits. Unlike yeast and mammals, in which only a single copy of each subunit is encoded,Arabidopsis encodes a multi-gene family for each subunit in its genome. Compared with the NF-Y of mammals or yeast, very little is known about plant NF-Y homologs. HereArabidopsis NF-YA subunits were isolated to determine whether they could form a hete-rotrimeric NF-Y complex with mammalian NF-YB and NF-YC. This resultant chimeric NF-Y complex had DNA binding ability to the same CCAAT sequences as those of the other life systems. Therefore, it is possible that plant NF-Y homologs might have biochemical characteristics similar to mammalian NF-Y, thereby suggesting its functional conservation among organisms.     

Identification,characterization and interaction of <Emphasis Type="Italic">HAP</Emphasis> family genes in rice

A HAP complex, which consists of three subunits, namely HAP2 (also called NF-YA or CBF-B), HAP3 (NF-YB/CBF-A) and HAP5 (NF-YC/CBF-C), binds to CCAAT sequences in a promoter to control the expression of target genes. We identified 10 HAP2 genes, 11 HAP3 genes and 7 HAP5 genes in the rice genome. All the three HAP family genes encode a protein with a conserved domain in each family and various non-conserved regions in both length and amino acid sequence. These genes showed various expression patterns depending on genes, and various combinations of overlapped expression of the HAP2, HAP3 and HAP5 genes were observed. Furthermore, protein interaction analyses showed interaction of OsHAP3A, a ubiquitously expressed HAP3 subunit of rice, with specific members of HAP5. These results indicate that the formation of specific complex with various HAP subunits combinations can be achieved by both tissue specific expression of three subunit genes and specific interaction of three subunit proteins. This may suggest that the HAP complexes may control various aspects of rice growth and development through tissue specific expression and complex formation of three subunit members. Nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession numbers AB288027 to AB288048 and BR000373 to BR000375.     

Requirement for down-regulation of the CCAAT-binding activity of the NF-Y transcription factor during skeletal muscle differentiation

NF-Y is composed of three subunits, NF-YA, NF-YB, and NF-YC, all required for DNA binding. All subunits are expressed in proliferating skeletal muscle cells, whereas NF-YA alone is undetectable in terminally differentiated cells in vitro. By immunohistochemistry, we show that the NF-YA protein is not expressed in the nuclei of skeletal and cardiac muscle cells in vivo. By chromatin immunoprecipitation experiments, we demonstrate herein that NF-Y does not bind to the CCAAT boxes of target promoters in differentiated muscle cells. Consistent with this, the activity of these promoters is down-regulated in differentiated muscle cells. Finally, forced expression of the NF-YA protein in cells committed to differentiate leads to an impairment in the down-regulation of cyclin A, cyclin B1, and cdk1 expression and is accompanied by a delay in myogenin expression. Thus, our results indicate that the suppression of NF-Y function is of crucial importance for the inhibition of several cell cycle genes and the induction of the early muscle-specific program in postmitotic muscle cells.     

Dissection of the NF-Y transcriptional activation potential.

NF-Y is a trimeric CCAAT-binding factor with histone fold subunits (NF-YB/NF-YC) and bipartite activation domains located on NF-YA and NF-YC. We reconstituted the NF-Y activation potential in vivo with GAL4 DBD fusions. In the GAL4-YA configuration, activation requires co-expression of the three subunits; with GAL4-YB and GAL4-YC, transfections of the histone fold partners are sufficient, provided that the Q-rich domain of NF-YC is present. Combinations of mutants indicate that the Q-rich domains of NF-YA and NF-YC are redundant in the trimeric complex. Glutamines 101 and 102 of NF-YA are required for activity. We assayed NF-Y on different promoter targets, containing single or multiple GAL4 sites: whereas on a single site NF-Y is nearly as powerful as VP16, on multiple sites neither synergistic nor additive effects are observed. NF-Y activates TATA and Inr core elements and the overall potency is in the same range as other Q-rich and Pro-rich activation domains. These results represent the first in vivo evidence of subunit interactions studies and further support the hypothesis that NF-Y is a general promoter organizer rather than a brute activator.     

大麦NF-YC基因鉴定及在盐胁迫下的表达分析

核因子Y (NF-Y)是由NF-YA、NF-YB和NF-YC三个亚基组成的一类真核细胞转录因子,主要参与植物生长发育调控和非生物胁迫信号传递。该研究利用生物信息学方法解析了大麦(Hordeum vulgare) NF-YC基因家族功能。首先,基于大麦基因组数据库鉴定出11个HvNF-YC成员,分布在除第2号染色体以外的其余6条染色体上,内含子0–5个。系统进化分析显示,大麦、拟南芥(Arabidopsis thaliana)和水稻(Oryza sativa) NF-YC基因家族成员可分为5个亚家族。基因复制分析显示, 6个HvNF-YC基因存在片段复制,3个HvNF-YC基因存在串联复制。启动子顺式作用元件分析显示,大多数HvNF-YC基因启动子含有与非生物胁迫及激素响应相关的顺式作用元件。对HvNF-YC家族成员在不同组织不同时期的表达模式分析表明,不同成员的时空表达存在明显差异,其中HvNF-YC9和HvNF-YC11可能在籽粒发育初期发挥重要作用。通过分析耐盐型和盐敏感型大麦品种根和叶中HvNF-YC表达量变化,发现HvNF-YC3、HvNF-YC6和HvNF-YC10主要在盐胁...