Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Tryptophan-2,3-dioxygenase (TDO) inhibition ameliorates neurodegeneration by modulation of kynurenine pathway metabolites

doi: 10.1073/pnas.1604453113

Figure Lengend Snippet: QUIN exacerbates neurodegeneration in HD flies and overexpression of hKAT is neuroprotective via increased KYNA levels. (A) QUIN levels in WT and HTT93Q-expressing flies. QUIN is detected in flies fed with 0.5 mg/mL of QUIN, but was not measurable in untreated flies. n = 3–5 flies per treatment, ***P < 0.001. (B) HTT93Q and cn−/− HTT93Q flies fed QUIN exhibit increased rhabdomere degeneration compared with untreated flies. Neuroprotection conferred by the cn mutation is abolished by QUIN feeding. n = 11–12 per treatment, **P < 0.01, ***P < 0.001. (C) Panneuronal overexpression of hKAT in a WT background causes an increase in KYNA production compared with controls at both posteclosion ages tested. n = 3–5 per genotype, ***P < 0.001. (D) HTT93Q flies with panneuronal overexpression of hKAT show a significant reduction in the 3-HK/KYNA ratio. The transgene control used in this experiment was a transgenic Drosophila line expressing an empty pJFRC2 vector. n = 4–5 per condition, **P < 0.01, ***P < 0.001. (E) Overexpression of hKAT is neuroprotective in HTT93Q flies at both posteclosion ages tested. n = 9–13 flies per condition, ***P < 0.001. (F) Overexpression of hKAT ameliorates the eclosion phenotype observed in HTT93Q flies. Transgene control + Htt93Q flies: n = 1084; hKAT + Htt93Q flies: n = 1,010, ***P < 0.001; ns, not significant. Data are the mean ± SEM (one-way ANOVA with Newman–Keuls post hoc test).

Article Snippet: The gene encoding kynurenine aminotransferase (hKAT) was amplified from a human fetal cDNA library ( 54 ) and cloned into the pJFRC2 vector ( 55 )—a gift from Gerald Rubin (Addgene plasmid no. 26214)—by standard methods.

Techniques: Over Expression, Expressing, Mutagenesis, Control, Transgenic Assay, Plasmid Preparation

Journal:

Article Title: The chvH Locus of Agrobacterium Encodes a Homologue of an Elongation Factor Involved in Protein Synthesis

doi: 10.1128/JB.183.1.36-45.2001

Figure Lengend Snippet: Bacterial strains and plasmids used in this study

Article Snippet: These concentrations were reduced by one-half for liquid medium. table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Strain or plasmid Description Reference or source Agrobacterium strains A348 A136(pTiA6) 21 A6007 A348 Pho − Sm r 11 A6880 A6007 chvH ::Tn phoA 12 At13000 A348 Δ chvH ::Ω fragment, contains pWT187kan This study At13001 At13000 without pWT187kan This study E. coli strains DH5α endA1 hsdR17 supE44 thi-1 recA1 gyrA96 relA1 Δ( argF-lacZYA ) U169 φ80d lacZ ΔM15 Gibco-BRL TG1 supE hsd Δ5 thi Δ( lac-proAB ) F′ ( traD36 proAB + lacI q lacZ ΔM15) U.S. Biochemical Corp. Plasmids pACL2 pRK7813 cosmid clone which complements A6880 12 pSW172 IncP broad-host-range vector with lac promoter 14 pSW213 IncP broad-host-range vector with lac promoter and lacI q 14 pTC110 pUCD2 Δ Pvu II- Eco RV 13 pBBR1MCS-4 Broad-host-range vector 32 pUC18 Multicopy vector 55 pUC19Ω Sma I Ω fragment in pUC19 10 pPR1068 pMAL-c2 derivative; Nde I at the start of MalE New England Biolabs pSP329 pTJS75 derivative with α-complementation group and multicloning site from pUC18, IncP, Tc r 11 pSP329Gm pSP329 derivative, IncP, Gm r D. M. Raineri pJQ200SK sacB suicide vector 44 pUCD2 IncW broad-host-range vector 16 pBSIIKS+.

Techniques: Plasmid Preparation