Journal: Nucleic Acids Research

Article Title: Evolutionary conservation supports ancient origin for Nudt16, a nuclear-localized, RNA-binding, RNA-decapping enzyme

doi: 10.1093/nar/gkn605

Figure Lengend Snippet: Amino acid alignment of putative Nudt16 orthologs from a sampling of the organisms found to have this protein. The alignment reveals three areas of highly conserved sequence, indicated by the green, red and blue boxes. A functional role based on structure for each conserved region is described in more detail in the text. To be designated as a Nudt16 homolog, sequences had to have the NUDIX domain (in red box) and the other two domains. Gray residues indicate identity; yellow highlighted residues show similarity. The accession number for each sequence used is listed in Supplementary Table 1.

Article Snippet: When arranged by degree of sequence divergence, the data indicated that the Syndesmos orthologs were grouped together as a distinct branch, diverging from the amniota branch of the Nudt16 tree ( ). is a schematic indicating the branches where Syndesmos (S) and Nudt16 (N) proteins have been found on a more simplified phylogenetic tree.

Techniques: Sampling, Sequencing, Functional Assay

Journal: Nucleic Acids Research

Article Title: Evolutionary conservation supports ancient origin for Nudt16, a nuclear-localized, RNA-binding, RNA-decapping enzyme

doi: 10.1093/nar/gkn605

Figure Lengend Snippet: A ‘standard’ phylogenetic tree. This tree illustrates the relative position of the organisms examined here and is provided as a quick reference for taxonomic relationships, based on the Tree of Life Web Project. ( A ) The Kingdom Animalia/Metazoa was searched. Only those branches for organisms containing a Nudt16 ortholog are shown. ( B ) An expansion of the vertebrate branch reveals the conservation of Nudt16 and appearance of Syndesmos. The ‘N’ indicates branches with organisms containing Nudt16 orthologs, ‘S’ indicates branches containing Syndesmos and ‘asterisk’ indicates the likely gene duplication event; acknowledging the possibility that it may have occurred earlier with high rates of subsequent gene loss. Sequences ‘below’ the ‘asterisk’ have only the Nudt16 protein.

Article Snippet: When arranged by degree of sequence divergence, the data indicated that the Syndesmos orthologs were grouped together as a distinct branch, diverging from the amniota branch of the Nudt16 tree ( ). is a schematic indicating the branches where Syndesmos (S) and Nudt16 (N) proteins have been found on a more simplified phylogenetic tree.

Techniques:

Journal: Nucleic Acids Research

Article Title: Evolutionary conservation supports ancient origin for Nudt16, a nuclear-localized, RNA-binding, RNA-decapping enzyme

doi: 10.1093/nar/gkn605

Figure Lengend Snippet: Insect Nudt16 protein is active for decapping RNA. Cap-labeled U8 RNA was incubated in the presence of buffer, Mn 2+ and protein (as indicated) for 30 min at 37°C (reaction indicated below). Samples of the reactions were spotted on a TLC, which was developed and then visualized on a phosphorimager. The Xenopus (X29/X-Nudt16) protein, present as a positive control, released m 7 GDP from the cap-labeled RNA. The two human proteins, which varied by an unrelated amino terminal extension, were both efficient at decapping the U8 RNA. Insect (sharpshooter) Nudt16 protein hydrolyzed the RNA to release the m 7 GPD cap comigrating with that cleaved by the other orthologs. The human H-Syndesmos protein displays no decapping activity.

Article Snippet: When arranged by degree of sequence divergence, the data indicated that the Syndesmos orthologs were grouped together as a distinct branch, diverging from the amniota branch of the Nudt16 tree ( ). is a schematic indicating the branches where Syndesmos (S) and Nudt16 (N) proteins have been found on a more simplified phylogenetic tree.

Techniques: Labeling, Incubation, Positive Control, Activity Assay

Journal: Nucleic Acids Research

Article Title: Evolutionary conservation supports ancient origin for Nudt16, a nuclear-localized, RNA-binding, RNA-decapping enzyme

doi: 10.1093/nar/gkn605

Figure Lengend Snippet: Nudt16 and Syndesmos are closely related members of a gene family. ( A) Alignment of a portion of the Syndesmos protein from 17 organisms (denoted by yellow box on right edge) and the corresponding region of Nudt16 from 10 organisms (marked with the blue box on right edge). All Syndesmos orthologs have a repeated glycine/leucine sequence (orange box) in place of the glutamic acid residues (heavy red box) required for catalysis in the NUDIX domain (red box). The green box is the conserved region in Nudt16 proteins, equivalent to the green box in . Syndesmos paralogs have all three conserved regions but lack a functional NUDIX domain. The central parts of the proteins are aligned here. The accession numbers are in Supplementary Table 1. ( B ) Genomic organization of the orthologs in human. Dashed boxes indicate the transcription unit, solid boxes are exons, horizontal lines are introns and the conserved domains are color coded as per . Note Nudt16 has a longer 5′-UTR and 3′-UTR than Syndesmos.

Article Snippet: When arranged by degree of sequence divergence, the data indicated that the Syndesmos orthologs were grouped together as a distinct branch, diverging from the amniota branch of the Nudt16 tree ( ). is a schematic indicating the branches where Syndesmos (S) and Nudt16 (N) proteins have been found on a more simplified phylogenetic tree.

Techniques: Sequencing, Functional Assay

Journal: Nucleic Acids Research

Article Title: Evolutionary conservation supports ancient origin for Nudt16, a nuclear-localized, RNA-binding, RNA-decapping enzyme

doi: 10.1093/nar/gkn605

Figure Lengend Snippet: Structural comparison of paralogous proteins. ( A ) Molecular modeling of the orthologs and paralogs of Xenopus Nudt16p. The Xray crystal structure of Xenopus X29/Nudt16p (PDB: 2A8P) was used to model the human and sharpshooter Nudt16p proteins and human Syndesmos. Selected conserved residues were highlighted in the ribbon models to correctly align the models relative to each other. The blue strand (CVTHFY) is indicated with blue asterisks in . The black asterisk is a conserved Phe residue near the catalytic site, while the red asterisk is a second conserved Phe residue indicated in . The yellow line in the Xenopus protein is oriented across the dimer interface; the monomer above the line is oriented and positioned relative to the other three proteins. ( B ) Electrostatic charge potential of structures. Once the proteins were correctly oriented, electrostatic charge potential was calculated. Blue is positively charged surface and red is negatively charged. The yellow line in the Xenopus protein is oriented across the dimer interface as in (A).

Article Snippet: When arranged by degree of sequence divergence, the data indicated that the Syndesmos orthologs were grouped together as a distinct branch, diverging from the amniota branch of the Nudt16 tree ( ). is a schematic indicating the branches where Syndesmos (S) and Nudt16 (N) proteins have been found on a more simplified phylogenetic tree.

Techniques: Comparison, Residue