Chang-Cai Liu

Genome-wide Identification and Characterization of a Dehydrin Gene Family in Poplar (Populus trichocarpa)

Dehydrins (DHNs) define a complex group of stress inducible proteins characterized by the presence of one or more lysine-rich motifs. DHNs are present in multiple copies in the genome of plant species. Although genome-wide analysis of DHNs composition and chromosomal distribution has been conducted in herbaceous species, it remains unexplored in woody plants. Here, we report on the identification of ten genes encoding eleven putative DHN polypeptides in Populus. We document that DHN genes occur as duplicated blocks distributed over seven of the 19 poplar chromosomes likely as a result of segmental and tandem duplication events. Based on conserved motifs, poplar DHNs were assigned to four subgroups with the Kn subgroup being the most frequent. One putative DHN polypeptide (PtrDHN-10) with a SKS arrangement could originate from a recombination between SKn and KnS genes. In silico analysis of microarray data showed that in unstressed poplar, DHN genes are expressed in all vegetative tissues except for mature leaves. This exhaustive survey of DHN genes in poplar provides important information that will assist future studies on their functional role in poplar.

Identification and analysis of phosphorylation status of proteins in dormant terminal buds of poplar

BackgroundAlthough there has been considerable progress made towards understanding the molecular mechanisms of bud dormancy, the roles of protein phosphorylation in the process of dormancy regulation in woody plants remain unclear.ResultsWe used mass spectrometry combined with TiO2 phosphopeptide-enrichment strategies to investigate the phosphoproteome of dormant terminal buds (DTBs) in poplar (Populus simonii × P. nigra). There were 161 unique phosphorylated sites in 161 phosphopeptides from 151 proteins; 141 proteins have orthologs in Arabidopsis, and 10 proteins are unique to poplar. Only 34 sites in proteins in poplar did not match well with the equivalent phosphorylation sites of their orthologs in Arabidopsis, indicating that regulatory mechanisms are well conserved between poplar and Arabidopsis. Further functional classifications showed that most of these phosphoproteins were involved in binding and catalytic activity. Extraction of the phosphorylation motif using Motif-X indicated that proline-directed kinases are a major kinase group involved in protein phosphorylation in dormant poplar tissues.ConclusionsThis study provides evidence about the significance of protein phosphorylation during dormancy, and will be useful for similar studies on other woody plants.

Phosphoproteomic identification and phylogenetic analysis of ribosomal P-proteins in Populus dormant terminal buds

To better understand the role that reversible phosphorylation plays in woody plant ribosomal P-protein function, we initiated a phosphoproteomic investigation of P-proteins from Populus dormant terminal buds. Using gel-free (in-solution) protein digestion and phosphopeptide enrichment combined with a nanoUPLC–ESI–MS/MS strategy, we identified six phosphorylation sites on eight P-proteins from Populus dormant terminal buds. Among these, six Ser sites and one Thr site were identified in the highly conserved C-terminal region of eight P-proteins of various P-protein subfamilies, including two P0, two P1, three P2 and one P3 protein. Among these, the Thr site was shown to be novel and has not been identified in any other organisms. Sequence analysis indicated that the phosphothreonine sites identified in the C-terminus of Ptr RPP2A exclusively occurred in woody species of Populus, etc. The identified phosphopeptides shared a common phosphorylation motif of (S/T)XX(D/E) and may be phosphorylated in vivo by casein kinase 2 as suggested by using Scansite analysis. Furthermore, phylogenetic analysis suggested that divergence of P2 also occurred in Populus, including type I and type II. To the best of our knowledge, this is the first systematic phosphoproteomic and phylogenetic analysis of P-proteins in woody plants, the results of which will provide a wealth of resources for future understanding and unraveling of the regulatory mechanisms of Populus P-protein phosphorylation during the maintenance of dormancy.

Identification and Analysis of the Acetylated Status of Poplar Proteins Reveals Analogous N-Terminal Protein Processing Mechanisms with Other Eukaryotes

Background The N-terminal protein processing mechanism (NPM) including N-terminal Met excision (NME) and N-terminal acetylation (Nα-acetylation) represents a common protein co-translational process of some eukaryotes. However, this NPM occurred in woody plants yet remains unknown. Methodology/Principal Findings To reveal the NPM in poplar, we investigated the Nα-acetylation status of poplar proteins during dormancy by combining tandem mass spectrometry with TiO2 enrichment of acetylated peptides. We identified 58 N-terminally acetylated (Nα-acetylated) proteins. Most proteins (47, >81%) are subjected to Nα-acetylation following the N-terminal removal of Met, indicating that Nα-acetylation and NME represent a common NPM of poplar proteins. Furthermore, we confirm that poplar shares the analogous NME and Nα-acetylation (NPM) to other eukaryotes according to analysis of N-terminal features of these acetylated proteins combined with genome-wide identification of the involving methionine aminopeptidases (MAPs) and N-terminal acetyltransferase (Nat) enzymes in poplar. The Nα-acetylated reactions and the involving enzymes of these poplar proteins are also identified based on those of yeast and human, as well as the subcellular location information of these poplar proteins. Conclusions/Significance This study represents the first extensive investigation of Nα-acetylation events in woody plants, the results of which will provide useful resources for future unraveling the regulatory mechanisms of Nα-acetylation of proteins in poplar.

In silico identification and characterization of N-Terminal acetyltransferase genes of poplar (Populus trichocarpa).

N-terminal acetyltransferase (Nats) complex is responsible for protein N-terminal acetylation (Nα-acetylation), which is one of the most common covalent modifications of eukaryotic proteins. Although genome-wide investigation and characterization of Nat catalytic subunits (CS) and auxiliary subunits (AS) have been conducted in yeast and humans they remain unexplored in plants. Here we report on the identification of eleven genes encoding eleven putative Nat CS polypeptides, and five genes encoding five putative Nat AS polypeptides in Populus. We document that the expansion of Nat CS genes occurs as duplicated blocks distributed across 10 of the 19 poplar chromosomes, likely only as a result of segmental duplication events. Based on phylogenetic analysis, poplar Nat CS were assigned to six subgroups, which corresponded well to the Nat CS types (CS of Nat A–F), being consistent with previous reports in humans and yeast. In silico analysis of microarray data showed that in the process of normal development of the poplar, their Nat CS and AS genes are commonly expressed at one relatively low level but share distinct tissue-specific expression patterns. This exhaustive survey of Nat genes in poplar provides important information to assist future studies on their functional role in poplar.

Genome-Wide Identification and in Silico Analysis of Poplar Peptide Deformylases

Peptide deformylases (PDF) behave as monomeric metal cation hydrolases for the removal of the N-formyl group (Fo). This is an essential step in the N-terminal Met excision (NME) that occurs in these proteins from eukaryotic mitochondria or chloroplasts. Although PDFs have been identified and their structure and function have been characterized in several herbaceous species, it remains as yet unexplored in poplar. Here, we report on the first identification of two genes (PtrPDF1A and PtrPDF1B) respectively encoding two putative PDF polypeptides in Populus trichocarpa by genome-wide investigation. One of them (XP_002300047.1) encoded by PtrPDF1B (XM_002300011.1) was truncated, and then revised into a complete sequence based on its ESTs support with high confidence. We document that the two PDF1s of Populus are evolutionarily divergent, likely as a result of independent duplicated events. Furthermore, in silico simulations demonstrated that PtrPDF1A and PtrPDF1B should act as similar PDF catalytic activities to their corresponding PDF orthologs in Arabidopsis. This result would be value of for further assessment of their biological activities in poplar, and further experiments are now required to confirm them.

The systematic characterization of poplar CK2α and its theoretical studies on phosphorylation of P-protein C-terminal domain

1 State Key Laboratory of Tree Genetics and Breeding (Northeast Forestry University), 26 Hexing Road, Harbin 150040, China. 2 School of Basic Medical Sciences, Jiamusi University, Jiamusi 154000, China. 3 Forestry college, Beihua University, Jilin 132013, China. 4 College of Forestry, Northeast Forestry University, Harbin 150040, China. 5 Laboratory for Chemical Defense and Microscale Analysis, P.O. Box 3, Zhijiang 443200, China. 6 Bureau of Garden and Park, 26 Shanhu Road, Qitaihe, 154600, China.