T Balla

Eunice Kennedy Shriver National Institute of Child Health and Human Development, Maryland, United States

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Publications (170)1079.61 Total impact

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    ABSTRACT: CDP-diacylglycerol Synthases (CDS)1 are critical enzymes that catalyze the formation of CDP-diacylglycerol (CDP-DAG) from phosphatidic acid (PA). Here we show in vitro that the two isoforms of human CDS, CDS1 and CDS2, show different acyl chain specificities for its lipid substrate. CDS2 is selective for the acyl chains at the sn-1 and sn-2 positions, the most preferred species being 1-stearoyl-2-arachidonoyl-sn-phosphatidic acid. CDS1, conversely, shows no particular substrate specificity, displaying similar activities for almost all substrates tested. Additionally, we show that inhibition of CDS2 by phosphatidylinositol is also acyl chain dependent, with the greatest inhibition seen with the 1-stearoyl-2-arachidonoyl species. CDS1 shows no acyl chain dependent inhibition. Both CDS1 and CDS2 are inhibited by their anionic phospholipid end products, with phosphatidylinositol-(4,5)-bisphosphate showing the greatest inhibition. Our results indicate that CDS1 and CDS2 could create different CDP-DAG pools that may serve to enrich different phospholipid species with specific acyl chains.
    Biochemistry 11/2014; · 3.38 Impact Factor
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    ABSTRACT: The yeast Efr3p protein is a major regulator of the Stt4p phosphatidylinositol 4-kinase at ER-PM contact sites. Its mutant fly homologue, Rbo displays diminishing light responses attributed to progressively impaired PLC signaling. Here we find that Efr3s play a role in maintaining responsiveness to angiotensin II (AngII) receptors. RNAi-mediated depletion of EFR3A and EFR3B impaired the sustained phase of cytosolic Ca(2+) response to high concentration of AngII in HEK293 cells expressing the wild type but not a truncated AT1a receptor, missing the phosphorylation sites. Efr3 depletion had minimal effect on the recovery of plasma membrane phosphoinositides during stimulation, and AT1 receptors still underwent ligand-induced internalization. A higher level of basal receptor phosphorylation and a larger response was observed after stimulation. Moreover, Gq activation more rapidly desensitized after AngII stimulation in Efr3 downregulated cells. Similar but smaller effect of EFR3 depletion was observed on the desensitization of the cAMP response after isoproterenol stimulation. These data suggest that mammalian Efr3s contribute to the control of the phosphorylation state and hence desensitization of AT1a receptors and could affect GPCR responsiveness in higher eukaryotes.
    Journal of cell science. 11/2014;
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    ABSTRACT: Phosphoinositides are a class of phospholipids generated by the action of phosphoinositide kinases with key regulatory functions in eukaryotic cells. Here, we present the atomic structure of phosphatidylinositol 4-kinase type IIα (PI4K IIα), in complex with ATP solved by X-ray crystallography at 2.8 Å resolution. The structure revealed a non-typical kinase fold that could be divided into N- and C-lobes with the ATP binding groove located in between. Surprisingly, a second ATP was found in a lateral hydrophobic pocket of the C-lobe. Molecular simulations and mutagenesis analysis revealed the membrane binding mode and the putative function of the hydrophobic pocket. Taken together, our results suggest a mechanism of PI4K IIα recruitment, regulation, and function at the membrane.
    EMBO Reports 10/2014; 15(10):1085-1092. · 7.19 Impact Factor
  • Gerald R V Hammond, Tamas Balla
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    ABSTRACT: The inositol lipids play many essential roles in eukaryotic physiology, although the action has usually focused on the special properties of their headgroup. Now, a study by Clark et al (2014) re-focuses attention on the hydrophobic lipid tails, showing that these too exhibit unique biochemical properties—and are also likely to play a fundamental role in the biology of the lipid.
    The EMBO Journal 09/2014; · 9.82 Impact Factor
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    ABSTRACT: Specificity of membrane fusion in vesicular trafficking is dependent on proper subcellular distribution of soluble N-ethylmaleimide sensitive factor attachment protein receptors (SNAREs). Although SNARE complexes are fairly promiscuous in vitro, significant specificity is achieved in cells due to spatial segregation and shielding of SNARE motifs prior to association with cognate Q-SNAREs. In this study we identified phosphatidylinositol 4-kinase IIα (PI4K2A) as a binding partner of vesicle-associated membrane protein 3 (VAMP3), a small R-SNARE involved in recycling and retrograde transport, and found that the two proteins co-reside on tubulo-vesicular endosomes. PI4K2A knockdown inhibited VAMP3 trafficking to perinuclear membranes and impaired the rate of VAMP3-mediated recycling of the transferrin receptor. Moreover, depletion of PI4K2A significantly decreased association of VAMP3 with its cognate Q-SNARE, Vti1a. Although binding of VAMP3 to PI4K2A did not require kinase activity, acute depletion of PtdIns4P on endosomes significantly delayed VAMP3 trafficking. Phospholipid modulation of SNARE function has been proposed based on in vitro studies and our study provides mechanistic evidence in support of these claims by identifying PI4K2A and PtdIns4P as regulators of an R-SNARE in intact cells.
    Journal of cell science. 07/2014;
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    ABSTRACT: Polyphosphoinositides are an important class of lipid that recruit specific effector proteins to organelle membranes. One member, phosphatidylinositol 4-phosphate (PtdIns4P) has been localized to Golgi membranes based on the distribution of lipid binding modules from PtdIns4P effector proteins. However, these probes may be biased by additional interactions with other Golgi-specific determinants. In this paper, we derive a new PtdIns4P biosensor using the PtdIns4P binding of SidM (P4M) domain of the secreted effector protein SidM from the bacterial pathogen Legionella pneumophila. PtdIns4P was necessary and sufficient for localization of P4M, which revealed pools of the lipid associated not only with the Golgi but also with the plasma membrane and Rab7-positive late endosomes/lysosomes. PtdIns4P distribution was determined by the localization and activities of both its anabolic and catabolic enzymes. Therefore, P4M reports a wider cellular distribution of PtdIns4P than previous probes and therefore will be valuable for dissecting the biological functions of PtdIns4P in its assorted membrane compartments.
    The Journal of Cell Biology 04/2014; · 10.82 Impact Factor
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    ABSTRACT: Vascular endothelial growth factor (VEGF) is a critical regulator of endothelial cell differentiation and vasculogenesis during both development and tumor vascularization. VEGF-165 is a major form that is secreted from the cells via a poorly characterized pathway. Here we used GFP and epitope-tagged VEGF-165 and found that its early trafficking between the ER and the Golgi requires the small GTP binding proteins, Sar1 and Arf1- and that its glycosylation in the Golgi compartment is necessary for efficient post Golgi transport and secretion from the cells. The relative temperature insensitivity of VEGF secretion and its Sar1 and Arf1 inhibitory profiles distinguished it from other cargoes using the "constitutive" secretory pathway. Prominent features of VEGF secretion were: the retention of the protein on the outer surface of the plasma membrane and the stimulation of its secretion by Ca(2+) and PKC. Importantly, shedding of VEGF-165 from the cell surface together with other membrane components appears to be a unique feature by which some of VEGF is delivered to the surroundings to exert its known biological actions. Understanding VEGF trafficking can reveal additional means by which tumor vascularization can be inhibited by pharmacological interventions.
    Molecular biology of the cell 02/2014; · 5.98 Impact Factor
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    ABSTRACT: Phosphatidylinositol 4-kinase type IIIα (PI4KA) is a host factor essential for Hepatitis C virus (HCV) replication and hence is a target for drug development. PI4KA has also been linked to ER-exit sites and generation of plasma membrane phosphoinositides. Here we developed highly specific and potent inhibitors of PI4KA and conditional knockout mice to study the importance of this enzyme in vitro and in vivo. Our studies showed that PI4KA is essential for the maintenance of plasma membrane PtdIns(4,5)P2 pools but only during strong stimulation of receptors coupled to PLC activation. Pharmacological blockade of PI4KA in adult animals leads to sudden death closely correlating with the drugs ability to induce PtdIns(4,5)P2 depletion after agonist stimulation. Genetic inactivation of PI4KA also leads to death, however, the cause in this case is due to severe intestinal necrosis. These studies highlight the risks of targeting PI4KA as an anti HCV strategy and also point to important distinctions between genetic and pharmacological studies when selecting host factors as putative therapeutic targets.
    Journal of Biological Chemistry 01/2014; · 4.65 Impact Factor
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    ABSTRACT: Glycosphingolipid (GSL) trafficking and metabolism are closely linked with cell fate determination, balancing signaling cascades controlling proliferation, apoptosis and autophagy. Altered cellular levels of complex glycosphingolipids often correlate with cancer progression, yet little is known about the principles that guide trafficking of these lipids.
    The FASEB Journal 01/2014; 28(1 Supplement). · 5.70 Impact Factor
  • Tamas Balla
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    ABSTRACT: Phosphoinositides (PIs) make up only a small fraction of cellular phospholipids, yet they control almost all aspects of a cell's life and death. These lipids gained tremendous research interest as plasma membrane signaling molecules when discovered in the 1970s and 1980s. Research in the last 15 years has added a wide range of biological processes regulated by PIs, turning these lipids into one of the most universal signaling entities in eukaryotic cells. PIs control organelle biology by regulating vesicular trafficking, but they also modulate lipid distribution and metabolism via their close relationship with lipid transfer proteins. PIs regulate ion channels, pumps, and transporters and control both endocytic and exocytic processes. The nuclear phosphoinositides have grown from being an epiphenomenon to a research area of its own. As expected from such pleiotropic regulators, derangements of phosphoinositide metabolism are responsible for a number of human diseases ranging from rare genetic disorders to the most common ones such as cancer, obesity, and diabetes. Moreover, it is increasingly evident that a number of infectious agents hijack the PI regulatory systems of host cells for their intracellular movements, replication, and assembly. As a result, PI converting enzymes began to be noticed by pharmaceutical companies as potential therapeutic targets. This review is an attempt to give an overview of this enormous research field focusing on major developments in diverse areas of basic science linked to cellular physiology and disease.
    Physiological Reviews 07/2013; 93(3):1019-137. · 30.17 Impact Factor
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    ABSTRACT: The BAR (Bin/Amphiphysin/Rvs) domain proteins arfaptin1 and arfaptin2 are localized to the trans-Golgi network (TGN) and, by virtue of their ability to sense and/or generate membrane curvature, could play an important role in the biogenesis of transport carriers. We report that arfaptins contain an amphipathic helix (AH) preceding the BAR domain, which is essential for their binding to phosphatidylinositol 4-phosphate (PI(4)P)-containing liposomes and the TGN of mammalian cells. The binding of arfaptin1, but not arfaptin2, to PI(4)P is regulated by protein kinase D (PKD) mediated phosphorylation at Ser100 within the AH. We also found that only arfaptin1 is required for the PKD-dependent trafficking of chromogranin A by the regulated secretory pathway. Altogether, these findings reveal the importance of PI(4)P and PKD in the recruitment of arfaptins at the TGN and their requirement in the events leading to the biogenesis of secretory storage granules.
    The EMBO Journal 05/2013; · 9.82 Impact Factor
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    ABSTRACT: The highly dynamic membranous network of eukaryotic cells allows spatial organization of biochemical reactions to suit the complex metabolic needs of the cell. The unique lipid composition of organelle membranes in the face of dynamic membrane activities assumes that lipid gradients are constantly generated and maintained. Important advances have been made in identifying specialized membrane compartments and lipid transfer mechanisms that are critical for generating and maintaining lipid gradients. Remarkably, one class of minor phospholipids - the phosphoinositides - is emerging as important regulators of these processes. Here, we summarize several lines of research that have led to our current understanding of the connection between phosphoinositides and the transport of structural lipids and offer some thoughts on general principles possibly governing these processes.
    Trends in cell biology 03/2013; · 12.12 Impact Factor
  • The FASEB Journal 01/2013; 27(1_MeetingAbstracts):1017.6. · 5.70 Impact Factor
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    ABSTRACT: A spectrin-based cytoskeleton is associated with endomembranes including the Golgi complex and cytoplasmic vesicles but its role remains poorly understood. Using new generated antibodies to specific peptide sequences of the human βIII spectrin, we here show its distribution in the Golgi complex, where it is enriched in the trans-Golgi and trans-Golgi network (TGN). The use of a drug-inducible enzymatic assay that deplete the Golgi-associated pool of PI4P as well as the expression of PH domains of Golgi proteins that specifically recognize this phosphoinositide both displaced βIII spectrin from the Golgi. However, the interference with actin dynamics using actin toxins did not affect the localization of βIII spectrin to Golgi membranes. Depletion of βIII spectrin using siRNA technology and the microinjection of anti-βIII spectrin antibodies into the cytoplasm lead to the fragmentation of the Golgi. At ultrastructural level, Golgi fragments showed swollen distal Golgi cisternae and vesicular structures. Using a variety of protein transport assays, we show that the ER-to-Golgi and post-Golgi protein transports were impaired in βIII spectrin-depleted cells. However, the internalization of the Shiga toxin subunit B to the ER was unaffected. We state that βIII spectrin constitutes a major skeletal component of distal Golgi compartments, where it is necessary to maintain its structural integrity and secretory activity, and unlike actin, PIP4 appears to be highly relevant for the association of βIII spectrin the Golgi complex.
    Journal of Biological Chemistry 12/2012; · 4.65 Impact Factor
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    ABSTRACT: The quantitatively minor phospholipid phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P(2)] fulfills many cellular functions in the plasma membrane (PM), whereas its synthetic precursor, phosphatidylinositol 4-phosphate (PI4P), has no assigned PM roles apart from PI(4,5)P(2) synthesis. We used a combination of pharmacological and chemical genetic approaches to probe the function of PM PI4P, most of which was not required for the synthesis or functions of PI(4,5)P(2). However, depletion of both lipids was required to prevent PM targeting of proteins that interact with acidic lipids or activation of the transient receptor potential vanilloid 1 cation channel. Therefore, PI4P contributes to the pool of polyanionic lipids that define plasma membrane identity and to some functions previously attributed specifically to PI(4,5)P(2), which may be fulfilled by a more general polyanionic lipid requirement.
    Science 06/2012; 337(6095):727-30. · 31.20 Impact Factor
  • Nihal Altan-Bonnet, Tamas Balla
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    ABSTRACT: Several RNA viruses have recently been shown to hijack members of the host phosphatidylinositol (PtdIns) 4-kinase (PI4K) family of enzymes. They use PI4K to generate membranes enriched in phosphatidylinositide 4-phosphate (PtdIns4P or PI4P) lipids, which can be used as replication platforms. Viral replication machinery is assembled on these platforms as a supramolecular complex and PtdIns4P lipids regulate viral RNA synthesis. This article highlights these recent studies on the regulation of viral RNA synthesis by PtdIns4P lipids. It explores the potential mechanisms by which PtdIns4P lipids can contribute to viral replication and discusses the therapeutic potential of developing antiviral molecules that target host PI4Ks as a form of panviral therapy.
    Trends in Biochemical Sciences 05/2012; 37(7):293-302. · 13.08 Impact Factor
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    ABSTRACT: Receptor endocytosis plays an important role in regulating the responsiveness of cells to specific ligands. Phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P(2)] has been shown to be crucial for endocytosis of some cell surface receptors, such as EGF and transferrin receptors, but its role in G-protein-coupled receptor internalization has not been investigated. By using luciferase-labeled type 1 angiotensin II (AT1R), type 2C serotonin (5HT2CR) or β(2) adrenergic (β2AR) receptors and fluorescently tagged proteins (β-arrestin-2, plasma-membrane-targeted Venus, Rab5) we were able to follow the sequence of molecular interactions along the endocytic route of the receptors in HEK293 cells using the highly sensitive method of bioluminescence resonance energy transfer and confocal microscopy. To study the role of plasma membrane PtdIns(4,5)P(2) in receptor endocytosis, we used our previously developed rapamycin-inducible heterodimerization system, in which the recruitment of a 5-phosphatase domain to the plasma membrane degrades PtdIns(4,5)P(2). Here we show that ligand-induced interaction of AT1, 5HT2C and β(2)A receptors with β-arrestin-2 was unaffected by PtdIns(4,5)P(2) depletion. However, trafficking of the receptors to Rab5-positive early endosomes was completely abolished in the absence of PtdIns(4,5)P(2). Remarkably, removal of the receptors from the plasma membrane was reduced but not eliminated after PtdIns(4,5)P(2) depletion. Under these conditions, stimulated AT1 receptors clustered along the plasma membrane, but did not enter the cells. Our data suggest that in the absence of PtdIns(4,5)P(2), these receptors move into clathrin-coated membrane structures, but these are not cleaved efficiently and hence cannot reach the early endosomal compartment.
    Journal of Cell Science 02/2012; 125(Pt 9):2185-97. · 5.88 Impact Factor
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    ABSTRACT: Gaucher disease is a lysosomal storage disorder caused by a defect in the degradation of glucosylceramide catalyzed by the lysosomal enzyme β-glucocerebrosidase (GBA). GBA reaches lysosomes via association with its receptor, lysosomal integral membrane protein type 2 (LIMP-2). We found that distinct phosphatidylinositol 4-kinases (PI4Ks) play important roles at multiple steps in the trafficking pathway of the LIMP-2/GBA complex. Acute depletion of phosphatidylinositol 4-phosphate in the Golgi caused accumulation of LIMP-2 in this compartment, and PI4KIIIβ was found to be responsible for controlling the exit of LIMP-2 from the Golgi. In contrast, depletion of PI4KIIα blocked trafficking at a post-Golgi compartment, leading to accumulation of LIMP-2 in enlarged endosomal vesicles. PI4KIIα depletion also caused secretion of missorted GBA into the medium, which was attenuated by limiting LIMP-2/GBA exit from the Golgi by PI4KIIIβ inhibitors. These studies identified PI4KIIIβ and PI4KIIα as important regulators of lysosomal delivery of GBA, revealing a new element of control to sphingolipid homeostasis by phosphoinositides.
    Molecular biology of the cell 02/2012; 23(8):1533-45. · 5.98 Impact Factor
  • The FASEB Journal 01/2012; 26(1_MeetingAbstracts):988.9. · 5.70 Impact Factor
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    ABSTRACT: Polyphosphoinositides are lipid signaling molecules generated from phosphatidylinositol (PtdIns) with critical roles in vesicular trafficking and signaling. It is poorly understood where PtdIns is located within cells and how it moves around between membranes. Here we identify a hitherto-unrecognized highly mobile membrane compartment as the site of PtdIns synthesis and a likely source of PtdIns of all membranes. We show that the PtdIns-synthesizing enzyme PIS associates with a rapidly moving compartment of ER origin that makes ample contacts with other membranes. In contrast, CDP-diacylglycerol synthases that provide PIS with its substrate reside in the tubular ER. Expression of a PtdIns-specific bacterial PLC generates diacylglycerol also in rapidly moving cytoplasmic objects. We propose a model in which PtdIns is synthesized in a highly mobile lipid distribution platform and is delivered to other membranes during multiple contacts by yet-to-be-defined lipid transfer mechanisms.
    Developmental Cell 11/2011; 21(5):813-24. · 12.86 Impact Factor

Publication Stats

8k Citations
1,079.61 Total Impact Points

Institutions

  • 2009–2014
    • Eunice Kennedy Shriver National Institute of Child Health and Human Development
      Maryland, United States
  • 2012
    • Rutgers, The State University of New Jersey
      • Federated Departments of Biological Sciences
      New Brunswick, NJ, United States
  • 1987–2012
    • National Institute of Child Health and Human Development
      Maryland, United States
    • National Institutes of Health
      • • Section on Molecular Signal Transduction
      • • Program in Developmental Neuroscience
      • • Section on Reproductive Endocrinology
      • • Laboratory of Cell and Developmental Signaling
      Bethesda, MD, United States
  • 1982–2012
    • Semmelweis University
      • Department of Physiology
      Budapest, Budapest fovaros, Hungary
  • 2011
    • University of Michigan
      • Department of Internal Medicine
      Ann Arbor, MI, United States
  • 2005–2010
    • Thomas Jefferson University
      • Department of Pathology, Anatomy & Cell Biology
      Philadelphia, PA, United States
  • 2001
    • Uniformed Services University of the Health Sciences
      • Department of Medicine
      Bethesda, MD, United States