Masur Auditorium | NIH Clinical Center, Building 10

Risky Business: Successes in the NIH Intramural Research Program

Transport of Large and Small Molecules across Bacterial Outer Membranes
Susan K. Buchanan, NIDDK

We use X-ray crystallography to study the structures and functions of integral membrane proteins in the outer membranes of bacteria. Bacteria require iron for survival, and they express transport systems dedicated to iron acquisition. Iron complexes are transported across the bacterial outer membrane using a transmembrane beta barrel protein coupled to an inner membrane protein (TonB) and proton motive force. We have been studying the questions of ligand specificity at the extracellular side of the transporter, and TonB recognition at the periplasmic side of the receptor. Exactly how these small molecules are transported through the barrel is not yet understood, despite the availability of several high resolution crystal structures. These same beta barrel proteins also serve as receptors/transporters for colicins, which are large proteins with bacteriocidal activity. This lecture will describe the differences between binding small molecules and large proteins to the same receptors, and will discuss how TonB and energy may be involved in the transport process(es). The knowledge gained from studying these systems may facilitate vaccine and drug design against Gram-negative bacterial pathogens.

RNAi and Epigenetic Mechanisms
Shiv Grewal, NCI

The sequencing of the genomes of several model organisms, and of the human genome, has led to new opportunities for exploring genome structure and function. Much attention has been focused on understanding how complex genomes are organized into distinct domains. In eukaryotic cells, genomic DNA is folded with histone and non-histone proteins in the form of chromatin. The recent discoveries of chromatin-remodeling machines, histone modifying activities, and/or modifications of DNA that can organize chromatin into accessible and inaccessible subdomains have revealed the existence of epigenetic mechanisms of genome regulation. Furthermore, there is growing realization that non-coding RNAs and the RNA interference (RNAi) mechanism play a fundamental role in epigenetic control of the genome and that small RNAs help guide epigenetic modifications to distinct chromosomal domains. A deeper understanding of the epigenetic mechanisms is crucial for the function of most, if not all, chromatin-templated processes with far-reaching consequences for human biology and diseases, including cancer and aging.

Cells with Nuclei Bent Out of Shape
Orna Cohen-Fix, NIDDK

The yeast nucleus, like that of higher eukaryotes, is a spherical organelle that is bound by a double membrane, the outer part of which is contiguous with the ER. The yeast nucleolus forms a crescent shape intranuclear structure that is associated with the nuclear membrane and caps the bulk of the chromosomal DNA. What controls the shape or size of intracellular organelles, such as the nucleus, is an important unanswered question. For example, certain types of cancer cells display an abnormal nuclear morphology, but the cause and effect of this phenotype are largely unknown. To gain a better understanding of how the yeast nuclear shape is determined, we studied the nuclear shape abnormality seen in the budding yeast spo7∆ mutant. In these cells, the nucleus exhibits a single protrusion that is devoid of DAPI-staining chromosomal DNA. The Spo7∆ protein is part of a phosphatase complex that represses phospholipids biosynthesis, suggesting that the nuclear protrusion seen in spo7∆ mutants is likely to be due to membrane expansion. Our studies reveal that this nuclear protrusion is restricted to the nuclear compartment occupied by the nucleolus, while the rest of the nucleus, containing the bulk of the DNA, retains its normal shape. Moreover, in diploid cells, which normally exhibit a single nucleolus, the absence of Spo7∆ often results in nuclei with two separate nucleoli, suggesting that the nuclear membrane plays an active role in maintaining the integrity of the nucleolus. We find that in spo7∆ mutants the peripheral ER membrane is also expanded, underscoring the fact that all ER membranes, with the exception of the membrane surrounding the bulk of the DNA, undergo expansion. Thus, the budding yeast nuclear membrane has distinct domains that are differentially susceptible to expansion. Moreover, these results suggest that there is a specialized mechanism that resists nuclear membrane expansion in the non-nucleolar domain. Possible mechanisms that may contribute to this process will be discussed.

Unraveling the Reactions of Nitric Oxide, Nitrite, and Hemoglobin in Human Physiology and Therapeutics
Mark Gladwin, NHLBI

Nitric oxide (NO) plays a fundamental role in maintaining normal vasomotor tone. Recent data implicate a critical function for hemoglobin and the erythrocyte in regulating the activity of NO in the vascular compartment. Intravascular hemolysis releases hemoglobin from the red blood cell into plasma (plasma cell-free plasma Hemoglobin), which is then able to scavenge endothelial derived NO 600-fold faster than erythrocytic hemoglobin, thereby disrupting NO homoestasis. This may lead to vasoconstriction, decreased blood flow, platelet activation, increased endothelin-1 expression (ET-1), and end-organ injury, and thus suggest a novel mechanism of disease for hereditary and acquired hemolytic conditions such as sickle cell disease and cardiopulmonary bypass.

In addition to providing an NO scavenging role in the physiological regulation of NO-dependent vasodilation, hemoglobin and the erythrocyte may deliver NO as the hemoglobin deoxygenates. While this process has previously been ascribed to S-nitrosated hemoglobin, recent data from our laboratories suggest that deoxygenated hemoglobin reduces nitrite to NO and vasodilates the human circulation along the physiological oxygen gradient. This newly described role of hemoglobin as a nitrite reductase is discussed in the context of blood flow regulation, oxygen sensing, and nitrite-based therapeutics.

Perspectives on the NIH Intramural Research Program: Past Progress, Future Promise
Elias A. Zerhouni, Director, NIH

Dr. Zerhouni will share his vision for the intramural research program, recognizing unique contributions it has made to NIH's research and training mission, and laying out future challenges as science becomes more complex and interdisciplinary. He will describe seminal experiences he has had as a research scientist, and underscore the critical importance of providing support to young investigators, especially in an era of unparalleled scientific opportunities. In addition, Dr. Zerhouni will share a new vision for clinical and translational sciences, and how the NIH intramural research program can contribute to that vision to advance 21st century medicine.