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

Article Title: δ-COP contains a helix C-terminal to its longin domain key to COPI dynamics and function

doi: 10.1073/pnas.1603544113

Figure Lengend Snippet: Yeast strains used in this study

Article Snippet: Ret2LD2α T158K ( BY4743 Spore ) MAT a his3 ∆ 1 leu2 ∆ 0 met15 ∆ 0 ura3 ∆ 0; YFR051c::kanMX4; p415 ret2LD2 α T158K This study Open in a separate window Yeast strains used in this study table ft1 table-wrap mode="anchored" t5 Table S2. caption a7 No. Plasmids Addgene ID Database ID Source 1. p415 CHI 1 73863 Z1280 This study 2. p415 CHI 2 73864 Z1281 This study 3. p415 CHI 3 73865 Z1283 This study 4. p415 CHI 4 73866 Z1284 This study 5. p415 CHI 5 73867 Z1285 This study 6. p415 CHI 6 73868 Z1286 This study 7. p415 CHI 7 73869 Z1287 This study 8. p415 CHI 8 73870 Z1288 This study 9. pRS303 Bt δCOP 73871 AE1540 This study 10. p415 Ret2 73872 AE1541 This study 11. p415 Ret2LD2α 73873 AE1518 This study 12. p415 δCLD2α 73874 AE1519 This study 13. p415 Ret2LD 73875 AE1520 This study 14. p415 δCLD 73876 AE1521 This study 15. p415 Ret2LD+δC2α 73877 AE1516 This study 16. p415 δCLD+Ret2-2α 73878 AE1523 This study 17. p415 CHI 9 73879 AF1551 This study 18. p415 CHI 10 73880 AF1553 This study 19. p415 CHI 12 73881 AF1557 This study 20. p415 CHI 13 73882 AH1658 This study 21. p415 CHI 14 73883 AH1660 This study 22. p415 CHI 15 73884 AH1662 This study 23. p415 Ret2 I145A 73885 AI1732 This study 24. p415 Ret2 I148A 73886 AI1733 This study 25. p415 Ret2 I149A 73887 AI1734 This study 26. p415 Ret2 I155A 73888 AI1735 This study 27. pGEX6P1- Mst7-Erp1CT 73920 This study 28. pGEX6P1-GCN4cc-Erp1CT 73921 This study 29. pGEX6P1- Mst7-Erp2CT 73922 This study 30. pGEX6P1-GCN4cc-Erp2CT 73923 This study 31. p415 Ret2LD2α Q146F 75259 This study 32. p415 Ret2LD2α N152T 75260 This study 33. p415 Ret2LD2α K153Q 75261 This study 34. p415 Ret2LD2α T158K 75262 This study 35. p416 PMP2-YFP-LRKRS — M650 14 36. p416 PMP2-YFP-KKXX — N654 14 37. p416 PMP2-YFP-NVRNRRK — R890 14 38. p416 PMP2-YFP-KKK — M643 14 39. p416 PMP2-YFP-AAXX — N655 14 40. p416 PMP2-YFP-Task3C44T — S901 27 41. pGEX6P1-MST27-LRKRS — F291 14 42. pGEX6P1-MST27-KKXX — O732 14 Open in a separate window Plasmids used in this study Secretion Assay.

Techniques:

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

Article Title: δ-COP contains a helix C-terminal to its longin domain key to COPI dynamics and function

doi: 10.1073/pnas.1603544113

Figure Lengend Snippet: Plasmids used in this study

Article Snippet: Ret2LD2α T158K ( BY4743 Spore ) MAT a his3 ∆ 1 leu2 ∆ 0 met15 ∆ 0 ura3 ∆ 0; YFR051c::kanMX4; p415 ret2LD2 α T158K This study Open in a separate window Yeast strains used in this study table ft1 table-wrap mode="anchored" t5 Table S2. caption a7 No. Plasmids Addgene ID Database ID Source 1. p415 CHI 1 73863 Z1280 This study 2. p415 CHI 2 73864 Z1281 This study 3. p415 CHI 3 73865 Z1283 This study 4. p415 CHI 4 73866 Z1284 This study 5. p415 CHI 5 73867 Z1285 This study 6. p415 CHI 6 73868 Z1286 This study 7. p415 CHI 7 73869 Z1287 This study 8. p415 CHI 8 73870 Z1288 This study 9. pRS303 Bt δCOP 73871 AE1540 This study 10. p415 Ret2 73872 AE1541 This study 11. p415 Ret2LD2α 73873 AE1518 This study 12. p415 δCLD2α 73874 AE1519 This study 13. p415 Ret2LD 73875 AE1520 This study 14. p415 δCLD 73876 AE1521 This study 15. p415 Ret2LD+δC2α 73877 AE1516 This study 16. p415 δCLD+Ret2-2α 73878 AE1523 This study 17. p415 CHI 9 73879 AF1551 This study 18. p415 CHI 10 73880 AF1553 This study 19. p415 CHI 12 73881 AF1557 This study 20. p415 CHI 13 73882 AH1658 This study 21. p415 CHI 14 73883 AH1660 This study 22. p415 CHI 15 73884 AH1662 This study 23. p415 Ret2 I145A 73885 AI1732 This study 24. p415 Ret2 I148A 73886 AI1733 This study 25. p415 Ret2 I149A 73887 AI1734 This study 26. p415 Ret2 I155A 73888 AI1735 This study 27. pGEX6P1- Mst7-Erp1CT 73920 This study 28. pGEX6P1-GCN4cc-Erp1CT 73921 This study 29. pGEX6P1- Mst7-Erp2CT 73922 This study 30. pGEX6P1-GCN4cc-Erp2CT 73923 This study 31. p415 Ret2LD2α Q146F 75259 This study 32. p415 Ret2LD2α N152T 75260 This study 33. p415 Ret2LD2α K153Q 75261 This study 34. p415 Ret2LD2α T158K 75262 This study 35. p416 PMP2-YFP-LRKRS — M650 14 36. p416 PMP2-YFP-KKXX — N654 14 37. p416 PMP2-YFP-NVRNRRK — R890 14 38. p416 PMP2-YFP-KKK — M643 14 39. p416 PMP2-YFP-AAXX — N655 14 40. p416 PMP2-YFP-Task3C44T — S901 27 41. pGEX6P1-MST27-LRKRS — F291 14 42. pGEX6P1-MST27-KKXX — O732 14 Open in a separate window Plasmids used in this study Secretion Assay.

Techniques:

Journal: Nature chemical biology

Article Title: Membrane editing with proximity labeling reveals regulators of lipid homeostasis

doi: 10.1038/s41589-025-02104-x

Figure Lengend Snippet: a , b , Quantification of miRFP-Nir2 (T59A mutant with lipid transfer activity disabled) co-localization with PA-fed membranes. Black horizontal bars indicate means and vertical error bars indicate standard deviations. Statistical significance was determined using two-sided Student’s t-test ( n = 10–16 cells with exact numbers provided in the Source Data, p = 8.6E-06 and 8.9E-07). c , Confocal images of HEK 293T cells stably expressing V5-tagged Nir2 (PITPNM1), PDZD8, ORP1L (OSBPL1A), full-length SCP2 (SCP-x), or the mature form of SCP2 (mSCP2). Cells were co-stained with α-LAMP1 and α-Calnexin antibodies as lysosome and ER markers. Shown are representative images from two independent experiments. d , e , Zoomed-in images of cells expressing V5-SCP-x ( d ) or V5-mSCP2 ( e ). V5-mSCP2 shows more cytosolic localization. Scale bars, 20 μm. f , Scheme of IMPACT labeling to quantify superPLD activity in live cells. g , h , Flow cytometry results of cells co-expressing superPLD targeted to the plasma membrane (PM; g ) or lysosomes (Lyso; h ) and the indicated lipid-transfer protein. mCherry signal (readout of superPLD expression level) and BODIPY signal (readout of PLD activity) are plotted. Wild-type cells expressing deadPLD were included as a negative control. Shown are representative plots from two independent experiments.

Article Snippet: Antibody and dilutions used for validation experiments ( – ) were the following: anti-LPIN1 polyclonal antibody (Proteintech, 27026–1-AP; 1:1,000), anti-LPIN2 monoclonal antibody (Santa Cruz Biotechnology, sc-514353; 1:100), anti-Nir2 polyclonal antibody (Proteintech, 26983–1-AP; 1:1,000), anti-PDZD8 polyclonal antibody (Proteintech, 25512–1-AP; 1:1,000), anti-SCP-x polyclonal antibody (Proteintech, 14397–1-AP; 1:1,000), anti-SCP-2 polyclonal antibody (Proteintech, 23006–1-AP; 1:1,000), anti-DGKD polyclonal antibody (Abcepta, AP8126b; 1:1,000), anti-DGKH polyclonal antibody (Proteintech, 13873–1-AP; 1:1,000) and anti-β-tubulin monoclonal antibody (Cell Signaling Technology, 86298).

Techniques: Mutagenesis, Activity Assay, Stable Transfection, Expressing, Staining, Labeling, Flow Cytometry, Clinical Proteomics, Membrane, Negative Control

Journal: Nature chemical biology

Article Title: Membrane editing with proximity labeling reveals regulators of lipid homeostasis

doi: 10.1038/s41589-025-02104-x

Figure Lengend Snippet: a , Western blot of LOVPLD-expressing HEK 293T cells treated with DsiRNA to deplete Nir2, SCP-x/SCP2, or PDZD8. DsiRNA #1 (colored red) was used in the subsequent experiments. , Quantification of ( a ), with relative levels compared to the control shown in parentheses. Plots are representative of two independent experiments. c , d , Expression levels of LOVPLD ( c ) and the PA probe GFP-PASS ( d ) in HEK 293T cells depleted with Nir2 (siNir2), SCP2 (siSCP2), or PDZD8 (siPDZD8), co-expressing PA probe and LOVPLD targeted to either plasma membrane (PM) or lysosomes (Lyso). Plots are representative of two independent experiments. e , f , Quantification of colocalization between LOVPLD and GFP-PASS shown in , . Black horizontal bars indicate means and vertical error bars indicate standard deviations. Statistical analysis was performed using one-way ANOVA followed by post-hoc Tukey-HSD test ( n = 14–19 cells with exact numbers provided in the Source Data, p = 0.01 and 0.6 for ( e ) and 0.03 and 0.9 for ( f ) for Control vs. siRNA samples). g , Western blot of p-S6K to measure mTOR activity in LTP-depleted cells expressing LOVPLD, with quantification presented in . LOVPLD was localized to plasma membrane (PM), lysosomes (Lyso), or endoplasmic reticulum (ER), and 30-min incubation with intermittent blue light illumination (470 nm, 500 ms per 5 s) was used to activate PA production by LOVPLD. β-actin was used as a loading control. h , Similar experiment as ( g ) but in LTP-depleted HEK 293T cells without LOVPLD activation. i , Quantification of ( h ). n = 4 (except for siPDZD8, where n = 2) biological replicates. Statistical analysis was performed using one-way ANOVA followed by post-hoc Tukey-HSD test ( p = 0.004, 0.01, and 0.07 for Control vs. siRNA samples). Black horizontal bars indicate means and vertical error bars indicate standard deviations.

Article Snippet: Antibody and dilutions used for validation experiments ( – ) were the following: anti-LPIN1 polyclonal antibody (Proteintech, 27026–1-AP; 1:1,000), anti-LPIN2 monoclonal antibody (Santa Cruz Biotechnology, sc-514353; 1:100), anti-Nir2 polyclonal antibody (Proteintech, 26983–1-AP; 1:1,000), anti-PDZD8 polyclonal antibody (Proteintech, 25512–1-AP; 1:1,000), anti-SCP-x polyclonal antibody (Proteintech, 14397–1-AP; 1:1,000), anti-SCP-2 polyclonal antibody (Proteintech, 23006–1-AP; 1:1,000), anti-DGKD polyclonal antibody (Abcepta, AP8126b; 1:1,000), anti-DGKH polyclonal antibody (Proteintech, 13873–1-AP; 1:1,000) and anti-β-tubulin monoclonal antibody (Cell Signaling Technology, 86298).

Techniques: Western Blot, Expressing, Control, Clinical Proteomics, Membrane, Activity Assay, Incubation, Activation Assay

Journal: International Journal of Molecular Sciences

Article Title: miR-24-3p and Body Mass Index as Type 2 Diabetes Risk Factors in Spanish Women 15 Years after Gestational Diabetes Mellitus Diagnosis

doi: 10.3390/ijms24021152

Figure Lengend Snippet: Boxplot diagram showing differential circulating miRNA expression profile in GDM women depending on their glycemic status after 15 years. ( a ) relative hsa-miR-1-3p expression, ( p = 0.385); ( b ) relative hsa-miR-24-3p expression ( p = 0.030); ( c ) relative hsa-miR-329-3p expression ( p = 0.596); ( d ) relative hsa-miR-543 expression ( p = 0.071). * p -value < 0.05 cel-miR-39-3p was used for miRNA expression normalization.

Article Snippet: hsa-miR-1-3p , 477820_mir , 5′-UGGAAUGUAAAGAAGUAUGUAU-3′.

Techniques: Expressing

Journal: Metallomics : integrated biometal science

Article Title: Differentiated Zn(II) binding affinities in animal, plant, and bacterial metallothioneins define their zinc buffering capacity at physiological pZn.

doi: 10.1093/mtomcs/mfad061

Figure Lengend Snippet: Fig. 8 Titration of metal-free BcrMT1B, XlMT, MacMT3, Ec-1, and TvMT with ZnSO 4 in the presence of ZnAF-2F. (A) 0.5 μM apo-MT and 2–3.5 μM ZnAF-2F (concentration varied depending on protein) in 50 mM Na + -HEPES, 100 mM NaCl, 500 μM TCEP. Orange line helps demonstrate titration end points. (B) Semilog plots of the same data are presented to visualize changes during titration. Blue lines help to determine the number of tight zinc sites. (C) Fluorescence responses were calibrated to obtain [Zn(II)] free concentrations and are shown as a pZn function of Zn(II)/apo-MT. Red lines demonstrate simulations based on calculated constants. −log K d values obtained from data fitting are presented in Table 3 (see section “Experimental procedures”). 47 Data are shown as means of n = 2 independent experiments + SD.

Article Snippet: All plasmids are deposited in Addgene under the following IDs: BcrMT1B (#200290), LIMT (#200291), SpMTA (#200292), XIMT (#200293), MacMT3 (#200294), OsMTI-1B (#200295), Ec-1 (#200296), PflQ2MT (#200297), SmtA (#200298), and TvMT (#200299).

Techniques: Titration, Concentration Assay, Fluorescence