Frederic BARD

Regulation of Membrane Trafficx

Profile

Frederic BARD
Lab Location: #5-12   Email: fbard@imcb.a-star.edu.sg   Tel: 65869585

Frederic Bard did his graduate work at Yale University, USA and at the Ecole Normale Superieure of Lyon, France where he obtained his PhD. He worked on the dynamics of the sealing zone, a unique actin cytoskeleton structure in osteoclasts required for bone resorption. During his postdoctoral work at the University of California San Diego (2001-2006), he identified a collection of new genes essential for general protein secretion, the TANGO genes. After starting his group IMCB in 2006, he has established a genome-wide RNAi screening platform. The group has focused on membrane trafficking regulation, Golgi organisation and how signalling at the Golgi can control protein glycosylation.

Regulation of Membrane Traffic

The human cell is organized into multiple intracellular compartments. Compartmentalization controls many aspects of cellular physiology and has increased in complexity throughout evolution. Most cellular compartments are bound, like the whole cell itself, by lipid-based membranes and exchange material through the trafficking of membrane-bound structures. We wish to understand how this membrane traffic is regulated to mediate various cellular functions. One of the technologies we use to address these questions is RNA interference screening at the genomic scale, which allows us to identify novel key players in these processes.
We focus on two questions:

1) How trafficking regulation at the Golgi complex affects glycosylation in health and disease
2) How intracellular trafficking is exploited by pathogens and toxins

The Golgi apparatus

Legend: Human Hela cells stained for the nucleus (blue), the microtubules network (red) and the Golgi apparatus (green).

 

Research

Lab projects:

1) Regulation of glycosylation through membrane traffic

The Golgi apparatus in human cells is an example of extensive compartmentalization, with multiple compartments- the cisternae- stacked together and multiple stacks connected together into a network. This complex structure is responsible for the synthesis of the various glycans presented at the cell surface.
In a published report, we have described how the organization of the Golgi apparatus can be altered to enhance the O-GalNac glycosylation of proteins upon activation of the known proto-oncogene Src. Interestingly, enhanced O-GalNac glycosylation is a known hallmark of cancerous transformation, with the up-regulation of the Tn antigen. We are now exploring the physiological significance of this O-glycosylation up-regulation.


Activation of Src at the Golgi apparatus promotes redistribution of GalNAc-Ts to the ER. HPL staining (marking the subcellular localization of GalNAc-Ts activity) is redistributed from the Golgi to the ER upon activation of endogenous Src in HeLa cells through treatment with EGF (100 ng/ml) for 4 h. After EGF stimulation, HPL staining revealed that GalNAc-Ts are active both in the ER (from the apparition of novel diffuse HPL-stained structures that co-localize with ER markers) and the ER-Golgi intermediate compartment (ERGIC) (from the apparition of novel punctate HPL-stained structures that co-localize with ERGIC markers). Golgi membranes are stained using giantin antibodies. Nuclei are stained blue with Hoescht. The scale bar represents 10mm. 

Src activation results in increased O-glycosylation initiation.
A model for Src-induced Golgi-to-ER retrograde trafficking of GalNac-T enzymes. Under normal cellular conditions, GalNac-Ts are predominantly localized to the cis-face of the Golgi apparatus and initiation of O-glycosylation occurs in the Golgi apparatus. Upon activation of Src, GalNac-Ts are selectively trafficked to the ER. This increases the efficiency of O-glycosylation initiation and results in a greater density of GalNAc added onto mucin-like proteins (further O-glycan modifications are not included for sake of clarity).

To test if other mechanisms are also regulating Golgi organization, we have undertaken, in another project, a large-scale RNAi screen to understand the genetic basis of Golgi organization.

For this, we have developed an automated analysis of Golgi morphological parameters.

We next used large-scale RNAi screening using the IMCB RNAi screening facility, which is embedded within the lab.

2) Regulation of retrograde membrane traffic and hijacking by toxins

In the secretory pathway, most proteins are synthesized in the endoplasmic reticulum (ER), then transported in an anterograde fashion to the Golgi and onwards to the plasma membrane, the endosomes or the extracellular space. By contrast, few proteins follow the opposite route: after endocytosis, they percolate through endosomes, reach the Golgi apparatus then move on to the ER. Various clinically important proteins, such as the Ricin, Cholera, Shiga and Pseudomonas toxins follow this retrograde trafficking pathway. In a published report, we have characterized the human genes required for intoxication by two toxins, Ricin and Pseudomonas Exotoxin. Most of these genes are likely to be implicated in the intracellular retrograde traffic. Their discovery offers many new insights into the mechanisms of retrograde membrane traffic.

Hypothetical model depicting how PE and Ricin travel from extracellular space to the ER
with some identified genes highlighted

Staff

Department: Frederic BARD

Name: Hui Hui WONG

Designation: Research Fellow

Email: hhwong@imcb.a-star.edu.sg


Name: Joanne Zhi Hui CHIA

Designation: Senior Research Fellow

Email: zhchia@imcb.a-star.edu.sg


Name: Alexandre CHAUMET

Designation: Senior Research Fellow

Email: achaumet@imcb.a-star.edu.sg


Name: Keit Min THAM

Designation: Senior Research Officer

Email: kmtham@imcb.a-star.edu.sg


Name: Xavier LE GUEZENNEC

Designation: Senior Research Fellow

Email: lgxavier@imcb.a-star.edu.sg


Name: Sze Hwee SEET

Designation: Senior Research Officer

Email: shseet@imcb.a-star.edu.sg


Name: Anh Tuan NGUYEN

Designation: Research Fellow

Email: atnguyen@imcb.a-star.edu.sg


Name: Trinda Anne TING

Designation: Senior Research Officer

Email: tating@imcb.a-star.edu.sg


Name: Jeremy Changyu YEO

Designation: Research Fellow

Email: cyyeo@imcb.a-star.edu.sg


Name: Le Son TRAN

Designation: Research Fellow

Email: tranls@imcb.a-star.edu.sg


Name: Eleanor ABBOTT

Designation: SIPGA Student

Email: eleanora@student.imcb.a-star.edu.sg


Name: Poonam SHAH

Designation: ARAP Student

Email: poonams@student.imcb.a-star.edu.sg


Name: Belinda Yi Ling TAN

Designation: Research Officer

Email: tanylb@imcb.a-star.edu.sg


Name: Felicia Pei Ling TAY

Designation: Research Officer

Email: pltay@imcb.a-star.edu.sg


Publications

Recent Publications

Wu KX, Phuektes P, Kumar P, Goh GY, Moreau D, Chow VT, Bard F, Chu JJ.
Human genome-wide RNAi screen reveals host factors required for enterovirus 71 replication.
Nat Commun. 2016 Oct 17;7:13150. doi: 10.1038/ncomms13150.

Calvert ME, Ward AM, Wright GD, Bard F.
New developments and novel applications in high throughput and high content imaging.
Cytometry A. 2016 Aug;89(8):705-7. doi: 10.1002/cyto.a.22921. No abstract available.

Toh CX, Chan JW, Chong ZS, Wang HF, Guo HC, Satapathy S, Ma D, Goh GY, Khattar E, Yang L, Tergaonkar V, Chang YT, Collins JJ, Daley GQ, Wee KB, Farran CA, Li H, Lim YP, Bard FA, Loh YH.
RNAi Reveals Phase-Specific Global Regulators of Human Somatic Cell Reprogramming.
Cell Rep. 2016 Jun 21;15(12):2597-607. doi: 10.1016/j.celrep.2016.05.049.

Ackema KB, Prescianotto-Baschong C, Hench J, Wang SC, Chia ZH, Mergentaler H, Bard F, Frank S, Spang A.
Sar1, a Novel Regulator of ER-Mitochondrial Contact Sites.
PLoS One. 2016 Apr 21;11(4):e0154280. doi: 10.1371/journal.pone.0154280.

Chia J, Goh G, Bard F.
Short O-GalNAc glycans: regulation and role in tumor development and clinical perspectives.
Biochim Biophys Acta. 2016 Aug;1860(8):1623-39. doi: 10.1016/j.bbagen.2016.03.008. Review.

Bard F, Chia J.
Cracking the Glycome Encoder: Signaling, Trafficking, and Glycosylation.
Trends Cell Biol. 2016 May;26(5):379-88. doi: 10.1016/j.tcb.2015.12.004. Review.

Lee WY, Goh G, Chia J, Boey A, Gunko NV, Bard F.
The Ubiquitin Ligase CBLC Maintains the Network Organization of the Golgi Apparatus.
PLoS One. 2015 Sep 22;10(9):e0138789. doi: 10.1371/journal.pone.0138789.

Yang BX, El Farran CA, Guo HC, Yu T, Fang HT, Wang HF, Schlesinger S, Seah YF, Goh GY, Neo SP, Li Y, Lorincz MC, Tergaonkar V, Lim TM, Chen L, Gunaratne J, Collins JJ, Goff SP, Daley GQ, Li H, Bard FA, Loh YH.
Systematic identification of factors for provirus silencing in embryonic stem cells.
Cell. 2015 Sep 24;163(1):230-45. doi: 10.1016/j.cell.2015.08.037.

Chaumet A, Wright GD, Seet SH, Tham KM, Gounko NV, Bard F.
Nuclear envelope-associated endosomes deliver surface proteins to the nucleus.
Nat Commun. 2015 Sep 10;6:8218. doi: 10.1038/ncomms9218.

Wong HH, Kumar P, Tay FP, Moreau D, Liu DX, Bard F.
Genome-Wide Screen Reveals Valosin-Containing Protein Requirement for Coronavirus Exit from Endosomes.
J Virol. 2015 Nov;89(21):11116-28. doi: 10.1128/JVI.01360-15.

Gonzales KA, Liang H, Lim YS, Chan YS, Yeo JC, Tan CP, Gao B, Le B, Tan ZY, Low KY, Liou YC, Bard F, Ng HH.
Deterministic Restriction on Pluripotent State Dissolution by Cell-Cycle Pathways.
Cell. 2015 Jul 30;162(3):564-79. doi: 10.1016/j.cell.2015.07.001.

Goh GY, Bard FA.
RNAi screens for genes involved in Golgi glycosylation.
Methods Mol Biol. 2015;1270:411-26. doi: 10.1007/978-1-4939-2309-0_28.

Ackema KB, Hench J, Böckler S, Wang SC, Sauder U, Mergentaler H, Westermann B, Bard F, Frank S, Spang A.
The small GTPase Arf1 modulates mitochondrial morphology and function.
EMBO J. 2014 Nov 18;33(22):2659-75. doi: 10.15252/embj.201489039.

Yu J, Chia J, Canning CA, Jones CM, Bard FA, Virshup DM.
WLS retrograde transport to the endoplasmic reticulum during Wnt secretion.
Dev Cell. 2014 May 12;29(3):277-91. doi: 10.1016/j.devcel.2014.03.016.

Chia J, Tham KM, Gill DJ, Bard-Chapeau EA, Bard FA.
ERK8 is a negative regulator of O-GalNAc glycosylation and cell migration.
Elife. 2014 Mar 11;3:e01828. doi: 10.7554/eLife.01828.

Dejeans N, Manié S, Hetz C, Bard F, Hupp T, Agostinis P, Samali A, Chevet E.
Addicted to secrete - novel concepts and targets in cancer therapy.
Trends Mol Med. 2014 May;20(5):242-50. doi: 10.1016/j.molmed.2013.12.003.

Bard-Chapeau EA, Nguyen AT, Rust AG, Sayadi A, Lee P, Chua BQ, New LS, de Jong J, Ward JM, Chin CK, Chew V, Toh HC, Abastado JP, Benoukraf T, Soong R, Bard FA, Dupuy AJ, Johnson RL, Radda GK, Chan EC, Wessels LF, Adams DJ, Jenkins NA, Copeland NG.
Transposon mutagenesis identifies genes driving hepatocellular carcinoma in a chronic hepatitis B mouse model.
Nat Genet. 2014 Jan;46(1):24-32. doi: 10.1038/ng.2847.

Kumar P, Goh G, Wongphayak S, Moreau D, Bard F.
ScreenSifter: analysis and visualization of RNAi screening data.
BMC Bioinformatics. 2013 Oct 3;14:290. doi: 10.1186/1471-2105-14-290.

Gill DJ, Tham KM, Chia J, Wang SC, Steentoft C, Clausen H, Bard-Chapeau EA, Bard FA.
Initiation of GalNAc-type O-glycosylation in the endoplasmic reticulum promotes cancer cell invasiveness.
Proc Natl Acad Sci U S A. 2013 Aug 20;110(34):E3152-61. doi: 10.1073/pnas.1305269110.

Bard-Chapeau EA, Gunaratne J, Kumar P, Chua BQ, Muller J, Bard FA, Blackstock W, Copeland NG, Jenkins NA.
EVI1 oncoprotein interacts with a large and complex network of proteins and integrates signals through protein phosphorylation.
Proc Natl Acad Sci U S A. 2013 Jul 30;110(31):E2885-94. doi: 10.1073/pnas.1309310110.

Mitrovic S, Nogueira C, Cantero-Recasens G, Kiefer K, Fernández-Fernández JM, Popoff JF, Casano L, Bard FA, Gomez R, Valverde MA, Malhotra V.
TRPM5-mediated calcium uptake regulates mucin secretion from human colon goblet cells.
Elife. 2013 May 28;2:e00658. doi: 10.7554/eLife.00658.

Sayadi, A., Nguyen, A.-T., Bard, F. A., & Bard-Chapeau, E. A.
Zip14 expression induced by lipopolysaccharides in macrophages attenuates inflammatory response.
Inflammation research : official journal of the European Histamine Research Society ... [et al.] (2013) 62(2), 133–143. doi:10.1007/s00011-012-0559-y

Chia J, Goh G, Racine V, Ng S, Kumar P, Bard F.
RNAi screening reveals a large signaling network controlling the Golgi apparatus in human cells.
Mol Syst Biol. 2012;8:629. doi: 10.1038/msb.2012.59.

Sayadi A, Nguyen AT, Bard FA, Bard-Chapeau EA.
Zip14 expression induced by lipopolysaccharides in macrophages attenuates inflammatory response.
Inflamm Res. 2013 Feb;62(2):133-43. doi: 10.1007/s00011-012-0559-y.

Moreau D, Bard F.
 [Looking for an antidote against A/B toxins].
Med Sci (Paris). 2012 Jan;28(1):28-31. doi: 10.1051/medsci/2012281010.

Moreau D, Kumar P, Wang SC, Chaumet A, Chew SY, Chevalley H, Bard F.
Genome-wide RNAi screens identify genes required for Ricin and PE intoxications.
Dev Cell. 2011 Aug 16;21(2):231-44. doi: 10.1016/j.devcel.2011.06.014.

Tjhi WC, Lee KK, Hung T, Tsang IW, Ong YS, Bard F, Racine V.
Exploratory analysis of cell-based screening data for phenotype identification in drug-siRNA study.
Int J Comput Biol Drug Des. 2011;4(2):194-215. doi: 10.1504/IJCBDD.2011.041011.

Dimitri Moreau, Pankaj Kumar, Shyi Chyi Wang, Alexandre Chaumet, Shin Yi Chew, Hélène Chevalley, Frédéric Bard.
Genome-Wide RNAi Screens Identify Genes Required for Ricin and PE Intoxications.
Dev Cell (2011) vol. 21 (2) pp. 231-44.

Gill DJ, Clausen H, Bard F.
Location, location, location: new insights into O-GalNAc protein glycosylation.
Trends Cell Biol. 2011 Mar;21(3):149-58. doi: 10.1016/j.tcb.2010.11.004. Review.

Chia NY, Chan YS, Feng B, Lu X, Orlov YL, Moreau D, Kumar P, Yang L, Jiang J, Lau MS, Huss M, Soh BS, Kraus P, Li P, Lufkin T, Lim B, Clarke ND, Bard F, Ng HH.
A genome-wide RNAi screen reveals determinants of human embryonic stem cell identity.
Nature. 2010 Nov 11;468(7321):316-20. doi: 10.1038/nature09531.

Gill DJ, Chia J, Senewiratne J, Bard F.
Regulation of O-glycosylation through Golgi-to-ER relocation of initiation enzymes.
J Cell Biol. 2010 May 31;189(5):843-58. doi: 10.1083/jcb.201003055.

Saito K, Chen M,Bard F, Chen S, Zhou H, Woodley D, Polischuk R, Schekman R, Malhotra V.
TANGO1 facilitates cargo loading at endoplasmic reticulum exit sites.
Cell. 2009 Mar 6;136(5):891-902.

RR Bard F, Malhotra V
Transit from the Trans-Golgi Network to the plasma membrane.
Annu Rev Cell Dev Biol.
2006;22:439-55.

Bard F, Casano L, Mallabiabarrena A, Wallace E, Saito K, Kitayama H, Guizzinti G, Hu Y, DasGupta R, Perrimon N, Malhotra V. Functional genomics reveals new genes involved in protein secretion and Golgi organization.
Nature. 2006 Feb 2;439(7076):604-7

Yeaman C, Ayala MI, Wright JR, Bard F, Bossard C, Ang A, Maeda Y, Seufferlein T, Mellman I, Nelson WJ, Malhotra V.
Protein kinase D regulates basolateral membrane protein exit from trans-Golgi network.
Nat Cell Biol.
2004 Feb;6(2):106-12.

Saltel F, Destaing O, Bard F, Eichert D, Jurdic P.
Apatite-mediated actin dynamics in resorbing osteoclasts.
Mol Biol Cell.
2004 15, 5231-41.

Bard F, Mazelin L, Pechoux-Longin C, Malhotra V, Jurdic P.
Src regulates Golgi structure and KDEL receptor-dependent retrograde transport to the endoplasmic reticulum.
J Biol Chem.
2003 Nov 21;278(47):46601-6.

Destaing O, Saltel F, Geminard JC, Jurdic P, Bard F. Podosomes display actin turnover and dynamic self-organization in osteoclasts expressing actin-green fluorescent protein.
Mol Biol Cell. 2003 Feb;14(2):407-16.