Jun Yang

• PhD
• PostDoc Position at Rutgers, The State University of New Jersey

Box C/D snoRNAs are known to guide site-specific ribose methylation of ribosomal RNA. Here, we demonstrate a novel and unexpected role for box C/D snoRNAs in guiding 18S rRNA acetylation in yeast. Our results demonstrate, for the first time, that the acetylation of two cytosine residues in 18S rRNA catalyzed by Kre33 is guided by two orphan box C/D...

Ribosomes are large ribonucleoprotein complexes that are fundamental for protein synthesis. Ribosomes are ribozymes because their catalytic functions such as peptidyl transferase and peptidyl-tRNA hydrolysis depend on the rRNA. rRNA is a heterogeneous biopolymer comprising of at least 112 chemically modified residues that are believed to expand its...

Methylation of ribose sugars at the 2′-OH group is one of the major chemical modifications in rRNA, and is catalyzed by snoRNA directed C/D box snoRNPs. Previous biochemical and computational analyses of the C/D box snoRNAs have identified and mapped a large number of 2′-OH ribose methylations in rRNAs. In the present study, we systematically analy...

RNA contains various chemical modifications that expand its otherwise limited repertoire to mediate complex processes like translation and gene regulation. 25S rRNA of the large subunit of ribosome contains eight base methylations. Except for the methylation of uridine residues, methyltransferases for all other known base methylations have been rec...

Yeast 25S rRNA was reported to contain a single cytosine methylation (m5C). In the present study using a combination of RP-HPLC, mung bean nuclease assay and rRNA mutagenesis, we discovered that instead of one, yeast contains two m5C residues at position 2278 and 2870. Furthermore, we identified and characterized two putative methyltransferases, Rc...

Epitranscriptomic modifications play pivotal roles in regulating RNA stability, localization and function. Recently, glycosylation has also emerged as an RNA modification, though its functional implications remain unclear. Here we report that metabolic labelling with a N-azidoacetylgalactosamine-tetraacylated bioorthogonal probe in mammalian cells...

Biallelic mutations in the DCPS gene disrupting the decapping activity of the scavenger decapping protein DcpS, leads to neurodevelopmental deficiencies and intellectual disability. However, the molecular basis for the neurogenesis defects in these individuals remains unknown. Here we show that cells derived from individuals with a DCPS mutation ha...

Epitranscriptomic modifications play pivotal roles in regulating RNA function, encompassing base alterations and the addition of both canonical m7G and noncanonical nucleotide metabolite caps. Recently, the spectrum of modifications has extended to include glyco modification at the 5' cap or within the RNA. Despite this expansion, the functional im...

Accurate identification of NAD-capped RNAs is essential for delineating their generation and biological function. Previous transcriptome-wide methods used to classify NAD-capped RNAs in eukaryotes contain inherent limitations that have hindered the accurate identification of NAD caps from eukaryotic RNAs. In this study, we introduce two orthogonal...

Identification of metabolite caps including FAD on the 5 end of RNA has uncovered a previously unforeseen intersection between cellular metabolism and gene expression. To understand the function of FAD caps in cellular physiology, we characterised the proteins interacting with FAD caps in budding yeast. Here we demonstrate that highly conserved 5-3...

Cellular RNAs, both coding and noncoding, contain several chemical modifications. Both ribose sugars and nitrogenous bases are targeted for these chemical additions. These modifications are believed to expand the topological potential of RNA molecules by bringing chemical diversity to otherwise limited repertoire. Here, using ribosomal RNA of yeast...

Identification of metabolite caps including FAD on the 5ʹ end of RNA has uncovered a previously unforeseen intersection between cellular metabolism and gene expression. To understand the function of FAD caps in cellular physiology, we characterised the proteins interacting with FAD caps in budding yeast. Here we demonstrate that highly conserved 5ʹ...

The existence of non-canonical nicotinamide adenine diphosphate (NAD) 5′-end capped RNAs is now well established. Nevertheless, the biological function of this nucleotide metabolite cap remains elusive. Here, we show that the yeast Saccharomyces cerevisiae cytoplasmic 5′-end exoribonuclease Xrn1 is also a NAD cap decapping (deNADding) enzyme that r...

The existence of non-canonical nicotinamide adenine diphosphate (NAD) 5′-end capped RNAs is now well established. Nevertheless, the biological function of this nucleotide metabolite cap remains elusive. Here, we show that the yeast Saccharomyces cerevisiae cytoplasmic 5′-end exoribonuclease Xrn1 is also a NAD cap decapping (deNADding) enzyme that r...

SOD1 is known as the major cytoplasmic superoxide dismutase and an anticancer target. However, the role of SOD1 in cancer is not fully understood. Herein we describe the generation of an inducible Sod1 knockout in KRAS-driven NSCLC mouse model. Sod1 knockout markedly reduces tumor burden in vivo and blocks growth of KRAS mutant NSCLC cells in vitro...

We recently reported the presence of nicotinamide adenine dinucleotide (NAD)-capped RNAs in mammalian cells and a role for DXO and the Nudix hydrolase Nudt12 in decapping NAD-capped RNAs (de-NADding) in cells. Analysis of 5 caps has revealed that in addition to NAD, mammalian RNAs also contain other metabolite caps including flavin adenine dinucleo...

tRNAs from all domains of life contain modified nucleotides. However, even for the experimentally most thoroughly characterized model organism Escherichia coli not all tRNA modification enzymes are known. In particular, no enzyme has been found yet for introducing the acp3U modification at position 47 in the variable loop of eight E. coli tRNAs. He...

CRAC analysis revealed specific binding of Kre33 to leucine and serine tRNAs (A), 18S rRNA (B) and snoRNAs snR4 and snR45 (C). A) Overview of CRAC-results for Kre33 binding to serine and leucine tRNA species. The left y-axis shows the total number of times each nucleotide within an RNA fragment was mapped to the RNA sequence (x-axis); the right y-a...

Sedimentation profiles of snR4 and snR45 upon Kre33 depletion. Kre33 was depleted using a strain containing HA tagged Kre33 under galactose promoter (pGAL1::3HA-kre33) that was grown in YPGSR (yeast extract, peptone, galactose–sucrose–raffinose, 2% w/v each) to mid-log phase, washed in pre-warmed water and transferred to YPD for up to 12 h. We comp...

snR4 is involved in 18S rRNA acetylation at C1280. A) Overlaid RP-HPLC chromatograms of the nucleosides derived from fragments isolated using mung bean nuclease assay, containing ac4C1280 (oligo 34, black) and ac4C1773 (oligo 45, red) isolated from a strain lacking snR4 (Δsnr4). Loss of snR4 leads to complete loss of ac4C1280 without affecting ac4C...

Crosslinking of Nop56, Nop58 and Nop1 to snR4 and snR45. 2D structure models (top panel) based on phylogenetic comparison and DMS-modification for snR4 (A) and snR45 (B) showing Kre33 cross-linked sites (identified in the present study, yellow ovals), along with the binding regions (transparent) and cross-linking sites (opaque ovals) of Nop1 (purpl...

Physical interaction of snR4 and snR45 with 18S rRNA is indispensable for 18S rRNA acetylation. 2D models of snR4 (A) and snR45 (B) along with zoomed-in views (C and D) for mutated regions. The dotted red box in (B) indicate a mutation that interfered with stable expression of the snoRNA. See for further details the legend for Fig 5, F) RP-HPLC chr...

Hybrids containing snoRNA sequences. (PDF)

Strains, plasmids and oligonucleotides. (PDF)

Phylogenetic analysis of snR45. A) Alignment of snoRNAs homologous to snR45. Sequences were retrieved and aligned as described previously [4]. The guide sequences (GS1 and GS2) and the C/D and C′ motifs stand out due to their high level of conservation (deep blue shade). The non-canonical D′ motif is quite variable and assigned based on 2D-modellin...

Phylogenetic analysis of snR4 with alignment (A) and 2D-models (B) as described for S2 Fig. Note, Torulaspora delbrueckii snR45 could be modeled with a D′ box using a canonical sequence (CUGA, red ellipse) instead of the CUGU found in S. cerevisiae and closely related species. (PDF)

[This corrects the article DOI: 10.1371/journal.pone.0168873.].

Mapping of 2’O-methylguanosines (Gm) and 2’O-methylcytidines (Cm) of 18S rRNA using snoRNA deletion mutants. To corroborate the location of residues mapped by mung bean nuclease assay, snoRNA deletion mutants for the respective ribose methylations were used. A specific loss of peak corresponding to the respective modification in a deletion mutant v...

Oligonucleotides used in the present study for MBN protection assay. (PDF)

RP-HPLC chromatogram of a standard aqueous mixture of commercially available ribonucleosides. Identities of relevant peaks are mentioned in the table below. (PDF)

Mapping of base modifications 25S rRNA using rDNA point mutants. To validate the location of base modifications of 25S rRNA, rDNA point mutants were generated where the modified residues were point mutated in a plasmid-containing 35S rDNA transcribed under the control of native promoter in a strain with genomic rDNA deletion. Overlaid chromatograms...

25S rRNA fragments (isolated by MBN digestion) with their respective modifications profile. (PDF)

Yeast strains used in the present study. (PDF)

18S rRNA fragments (isolated by MBN digestion) with their respective modifications profile. (PDF)

An endo-polygalacturonase gene, pga1, was cloned from the acidophilic fungus Bispora sp. MEY-1 and expressed in Pichia pastoris. The 1455-bp full-length complementary DNA of pga1 encoded a 485-amino acid polypeptide (endo-PGA1) including a putative 21-residue signal peptide and a catalytic domain belonging to glycoside hydrolase family 28. Purified...

We cloned and sequenced a xylanase gene named xylD from the acidophilic fungus Bispora sp. MEY-1 and expressed the gene in Pichia pastoris. The 1,422-bp full-length complementary DNA fragment encoded a 457-amino acid xylanase with a calculated molecular mass of 49.8 kDa. The mature protein of XYLD showed high sequence similarity to both glycosyl hy...

A xylanase gene (xyl11B) was cloned from Bispora sp. MEY-1 and expressed in Pichia pastoris. xyl11B, with a 66-bp intron, encodes a mature protein of 219 residues with highest identity (57.1%) to the Trichoderma reesei xylanase of glycoside hydrolase family 11. The purified recombinant XYL11B was acidophilic, exhibiting maximum activity at pH 2.6 a...

Most reported microbial beta-1,3-1,4-glucanases belong to the glycoside hydrolase family 16. Here, we report a new acidic family 7 endo-beta-1,3-1,4-glucanase (Bgl7A) from the acidophilic fungus Bispora sp. MEY-1. The cDNA of Bgl7A was isolated and over-expressed in Pichia pastoris, with a yield of about 1,000 U ml(-1) in a 3.7-l fermentor. The pur...

A complete gene, xyl10C, encoding a thermophilic endo-1,4-beta-xylanase (XYL10C), was cloned from the acidophilic fungus Bispora sp. MEY-1 and expressed in Pichia pastoris. XYL10C shares highest nucleotide and amino acid sequence identities of 57.3 and 49.7%, respectively, with a putative xylanase from Aspergillus fumigatus Af293 of glycoside hydro...

Using degenerate polymerase chain reaction (PCR) and thermal asymmetric interlaced PCR, a 1,347-bp full-length complementary DNA fragment encompassing the gene man5A, which encodes a 429-amino acid beta-mannanase with a calculated mass of 46.8 kDa, was cloned from acidophilic Bispora sp. MEY-1. The deduced amino acid sequence (catalytic domain) dis...