dc3000 pks1 schyzosaccharomyces pombe fas2 ks neurospora crassa 74 or23 1a fas2 ks cyanophora paradoxa  (ATCC)

Journal: Fungal genetics reports

Article Title: The auxotrophic formate ( for ) mutant of Neurospora crassa has significantly delayed growth but a normal circadian clock

doi: 10.4148/1941-4765.2185

Figure Lengend Snippet: The canonical for allele carries a H223Y mutation in the PLP binding pocket of cytosolic serine hydroxymethyltransferase (SHMT). A) Amino acids sequence alignments of cytosolic or mitochondrial SHMT between Neurospora crassa and other organisms with the phylogenetic tree on the left. Conserved H223 is indicated by the black rectangle. B) Comparison between wild-type and mutated Alphafold-predicted structures of SHMT. Wild-type structure is shown in yellow. H223Y mutated structure is shown in blue.

Article Snippet: The filamentous fungus Neurospora crassa is a classical model organism for circadian research as well as other fields ( ; Roche et al . 2014 ).

Techniques: Mutagenesis, Binding Assay, Sequencing, Comparison

Journal: Nucleic Acids Research

Article Title: H3T11 phosphorylation by CKII is required for heterochromatin formation in Neurospora

doi: 10.1093/nar/gkae664

Figure Lengend Snippet: CKII deficiency leads to activation of cat-3 gene expression by disrupting its upstream 5-kb heterochromatin. Growth phenotypes of WT, CKII mutants and CKII complementary transformants on slants ( A ) and plates ( B ), respectively. ( C ) In-gel analysis of catalase activity in extracts from WT, CKII mutants, CKII complementary transformants and cat-3 KO strains. ( D ) Western blot analysis of CAT-3 protein in WT, CKII mutants, CKII complementary transformants and cat-3 KO strains. The membrane stained by Coomassie blue served as the loading control. ( E ) RT-qPCR assay analyzing the levels of cat-3 messenger RNA (mRNA) in WT, cka KO , ckb1 KO , ckb2 KO and ckb1 KO ckb2 KO strains. Error bars indicate standard deviation (SD; n = 3). *** P < 0.001. Unpaired Student’s t -test was used. ( F ) Schematic diagram displaying a 5-kb heterochromatin domain ( 5H-cat-3 domain) located between cat-3 (NCU00355) and NCU00354 genes on linkage group III of the N. crassa genome. Primer pairs from 1 to 8 under the schematic diagram indicate regions tested by ChIP-qPCR. ChIP assays with H3K9me3 and HP1 antibodies revealing the enrichment profiles of H3K9me3 ( G ) and HP1 ( H ) at the 5H-cat-3 domain in WT, cka KO and ckb1 KO ckb2 KO strains. The H3K9L and hpo KO strains were used as the negative controls in panels (G) and (H), respectively. Error bars indicate SD ( n = 3). *** P < 0.001; n.s., not significant. Unpaired Student’s t -test was used.

Article Snippet: Neurospora crassa total RNA was extracted with the TRIzol agent and treated with DNase I to digest genomic DNA, as described previously ( ): reverse transcription (RT) with Maxima H Minus reverse transcriptase (Thermo Fisher Scientific, #M1682) after quantification of 5 μg per RNA sample, and then amplification by real-time polymerase chain reaction (PCR) (7500; ABI).

Techniques: Activation Assay, Expressing, Activity Assay, Western Blot, Membrane, Staining, Control, Quantitative RT-PCR, Standard Deviation

Journal: Nucleic Acids Research

Article Title: H3T11 phosphorylation by CKII is required for heterochromatin formation in Neurospora

doi: 10.1093/nar/gkae664

Figure Lengend Snippet: CKII-dependent H3T11 phosphorylation is also required for the formation of the 50-kb heterochromatin region. ( A ) Schematic diagram displaying a 50-kb heterochromatin domain upstream of the met-8 gene on linkage group III of the N. crassa genome. Primer pairs from 1 to 7 under the schematic diagram indicate the regions tested by ChIP-qPCR. ( B ) ChIP assay with the CKA antibody displaying the binding profiles of CKA protein at the 50-kb heterochromatin domain in the WT, ckb1 KO ckb2 KO , cka D149A and H3T11A mutant strains; the cka KO mutant was used as the negative control. Error bars indicate SD ( n = 3). *** P < 0.001. Unpaired Student’s t -test was used. ( C ) ChIP assay with H3pT11 antibody showing the enrichment of H3pT11 at the 50-kb heterochromatin domain in the WT and cka KO strains. The H3T11A strain was used as the negative control. Error bars indicate SD ( n = 3). *** P < 0.001. Unpaired Student’s t -test was used. ChIP assays revealing the enrichment profiles of H3K9me3 ( D ) and HP1 ( E ) at the 50-kb heterochromatin in WT and cka KO strains. The H3K9L and hpo KO strains were used as the negative controls in panels (D) and (E), respectively. Error bars indicate SD ( n = 3). *** P < 0.001. Unpaired Student’s t -test was used. ChIP assays revealing the enrichment profiles of H3K9me3 ( F ) and HP1 ( G ) at the 50-kb heterochromatin in WT, H3T11A and H3T11E strains. The H3K9L and hpo KO strains were used as the negative controls in panels (F) and (G), respectively. Error bars indicate SD ( n = 3). *** P < 0.001. Unpaired Student’s t -test was used. ( H ) RT-qPCR assay analyzing the levels of met-8 mRNA in WT, cka KO , cka K61A , cka D149A , ckb1 KO ckb2 KO , H3T11A , H3T11E and dim-5 KO strains. Error bars indicate SD ( n = 3). *** P < 0.001. Unpaired Student’s t -test was used. ( I ) Western blot analysis of MET-8 protein in WT, cka KO , cka K61A , cka D149A , ckb1 KO ckb2 KO , H3T11A , H3T11E and dim-5 KO strains. The membrane stained by Coomassie blue served as the loading control.

Article Snippet: Neurospora crassa total RNA was extracted with the TRIzol agent and treated with DNase I to digest genomic DNA, as described previously ( ): reverse transcription (RT) with Maxima H Minus reverse transcriptase (Thermo Fisher Scientific, #M1682) after quantification of 5 μg per RNA sample, and then amplification by real-time polymerase chain reaction (PCR) (7500; ABI).

Techniques: Binding Assay, Mutagenesis, Negative Control, Quantitative RT-PCR, Western Blot, Membrane, Staining, Control

Journal: Applied microbiology and biotechnology

Article Title: Expression and purification of recombinant tung tree diacylglycerol acyltransferase 2

doi: 10.1007/s00253-012-3869-7

Figure Lengend Snippet: The evolutionary relationships among the 74 DGAT2s from 61 organisms were analyzed by phylogenetic analysis based on the Neighbor-Joining method of Saitou and Nei [54]. The name of each protein sequence consists of the initials of the organism followed by the assigned subfamily of DGATs in the databases and the GenBank accession number. The numbers in the parenthesis following DGAT names are the calculated distance values, which reflect the degree of divergence between all pairs of DGAT sequences analyzed. The abbreviations of the organisms are: Ac in AcDGAT1-EGC41804.1, Ajellomyces capsulatus; Ac in AcDGAT2-XP_003225477.1, Anolis carolinensis; Ac in AcDGAT2-XP_001273210, Aspergillus clavatus; Ag, Ashbya gossypii; Ao, Arthroderma otae; At, Arabidopsis thaliana; Bn, Brassica napus; Bt, Bos taurus; Ce, Caenorhabditis elegans; Ci in CiDGAT2-XP_002120879.1, Ciona intestinalis; Ci in CiDGAT2-XP_001240299, Coccidioides immitis; Cn, Cryptococcus neoformans: Cr, Chlamydomonas reinhardtii; Cv, Chlorella variabilis; Dd, Dictyostelium discoideum; Dr, Danio rerio; Ea, Euonymus alatus; Eo, Elaeis oleifera; Gm, Glycine max; Gz, Gibberella zeae; Ha, Helianthus annuus; Hs, Homo sapiens; Hv, Hordeum vulgare; Ip, Ictalurus punctatus; Lb, Laccaria bicolor; Md, Monodelphis domestica; Mm, Mus musculus; Mo, Magnaporthe oryzae; Mt, Medicago truncatula; Nc, Neurospora crassa; Nf, Neosartorya fischeri; Nv, Nematostella vectensis; Oa, Ovis aries; Oe, Olea europaea; Os, Oryza sativa; Ot, Ostreococcus tauri; Pa, Pongo abelii; Pm, Penicillium marneffei; Pn, Phaeosphaeria nodorum; Pp in PpDGAT2-EFA83646.1, Polysphondylium pallidum; Pp in PpDGAT2a-XP_001758758.1 and PpDGAT2b-XP_001777726.1, Physcomitrella patens; Ps, Picea sitchensis; Pt in PtDGAT2-XP_527842.2, Pan troglodytes; Pt in PtDGAT2-XP_002317635.1, Populus trichocarpa; Rc, Ricinus communis; Rn, Rattus norvegicus; Sb, Sorghum bicolor; Sc, Saccharomyces cerevisiae; Sm, Selaginella moellendorffii; Sp in SpDGAT2-AAQ89590.1, Spirodela polyrhiza; Sp in SpDGAT2-XP_001713160.1, Schizosaccharomyces pombe; Tc, Tribolium castaneum; Tg, Toxoplasma gondii; Ts, Talaromyces stipitatus; Um, Ustilago maydis; Ur, Umbelopsis ramanniana; Vf, Vernicia fordii; Vg, Vernonia galamensis; Vv, Vitis vinifera; Xt, Xenopus tropicalis; Zm, Zea mays. This analysis is updated with more DGAT2s from a previous analysis [32].

Article Snippet: The abbreviations of the organisms are: Ac in AcDGAT1-EGC41804.1, Ajellomyces capsulatus ; Ac in AcDGAT2-XP_003225477.1, Anolis carolinensis ; Ac in AcDGAT2-XP_001273210, Aspergillus clavatus ; Ag, Ashbya gossypii ; Ao, Arthroderma otae ; At, Arabidopsis thaliana ; Bn, Brassica napus ; Bt, Bos taurus ; Ce, Caenorhabditis elegans ; Ci in CiDGAT2-XP_002120879.1, Ciona intestinalis ; Ci in CiDGAT2-XP_001240299, Coccidioides immitis ; Cn, Cryptococcus neoformans : Cr, Chlamydomonas reinhardtii ; Cv, Chlorella variabilis ; Dd, Dictyostelium discoideum ; Dr, Danio rerio ; Ea, Euonymus alatus ; Eo, Elaeis oleifera ; Gm, Glycine max ; Gz, Gibberella zeae ; Ha, Helianthus annuus ; Hs, Homo sapiens ; Hv, Hordeum vulgare ; Ip, Ictalurus punctatus ; Lb, Laccaria bicolor ; Md, Monodelphis domestica ; Mm, Mus musculus ; Mo, Magnaporthe oryzae ; Mt, Medicago truncatula ; Nc, Neurospora crassa ; Nf, Neosartorya fischeri ; Nv, Nematostella vectensis ; Oa, Ovis aries ; Oe, Olea europaea ; Os, Oryza sativa ; Ot, Ostreococcus tauri ; Pa, Pongo abelii ; Pm, Penicillium marneffei ; Pn, Phaeosphaeria nodorum ; Pp in PpDGAT2-EFA83646.1, Polysphondylium pallidum ; Pp in PpDGAT2a-XP_001758758.1 and PpDGAT2b-XP_001777726.1, Physcomitrella patens ; Ps, Picea sitchensis ; Pt in PtDGAT2-XP_527842.2, Pan troglodytes ; Pt in PtDGAT2-XP_002317635.1, Populus trichocarpa ; Rc, Ricinus communis ; Rn, Rattus norvegicus ; Sb, Sorghum bicolor ; Sc, Saccharomyces cerevisiae ; Sm, Selaginella moellendorffii ; Sp in SpDGAT2-AAQ89590.1, Spirodela polyrhiza ; Sp in SpDGAT2-XP_001713160.1, Schizosaccharomyces pombe ; Tc, Tribolium castaneum ; Tg, Toxoplasma gondii ; Ts, Talaromyces stipitatus ; Um, Ustilago maydis ; Ur, Umbelopsis ramanniana ; Vf, Vernicia fordii ; Vg, Vernonia galamensis ; Vv, Vitis vinifera ; Xt, Xenopus tropicalis ; Zm, Zea mays .

Techniques: Sequencing