Commentary: Systems Biology and Its Relevance to Alcohol Research

Q. Max Guo, Ph.D., and Sam Zakhari, Ph.D.

Systems biology, a new scientific discipline, aims to study the behavior of a biological organization or process in order to understand the function of a dynamic system. This commentary will put into perspective topics discussed in this issue of Alcohol Research & Health, provide insight into why alcohol-induced disorders exemplify the kinds of conditions for which a systems biological approach would be fruitful, and discuss the opportunities and challenges facing alcohol researchers. KEY WORDS: Alcohol-induced disorders; alcohol research; biomedical research; systems biology; biological systems; mathematical modeling; genomics; epigenomics; transcriptomics; metabolomics; proteomics

ntil recently, most biologists’ emerging discipline that deals with, Alcohol Research & Health intend to efforts have been devoted to and takes advantage of, these enormous address. In this commentary, we will Ureducing complex biological amounts of data. Although scientists try to put the topics discussed in this systems to the properties of individual and engineers have applied the concept issue into perspective, provide views molecules. However, with the com­ of an integrated systemic approach for on the significance of systems biology pleted sequencing of the genomes of years, systems biology has only emerged as a new, distinct discipline to study 1 High-throughput genomics is the study of the structure humans, mice, rats, and many other and function of an organism’s complete genetic content, organisms, technological advances in complex biological systems in the past or genome, using technology that analyzes a large num­ the fields of high-throughput genomics1 several years. A database search using ber of genes at a time. See the glossary, p. 84, for descriptions of other technical terms used in this article. and functional genomics2 have gener­ the phrase “systems biology” in the ISI ated enormous amounts of information Web of Science has found only 3 pub­ 2 Functional genomics aims to discover, on a large scale, 3 the biological function of particular genes and to uncover on the properties of genes, RNAs, lications in 2001; in 2006, this number how sets of genes and their products work together in proteins, and metabolic products (i.e., had reached 575 (see figure 1). health and disease. An example includes gene expression metabolites) in an organism. The bil­ With the emergence of this power­ profiling. lions of data points generated by these ful new discipline, we are tempted to 3 RNA is mostly involved in the processes of translating high-throughput studies are far beyond ask the following questions: Is systems genetic information from DNA into proteins. Some non- coding RNAs also are directly involved in other biological the reach of reductionist approaches. biology suitable for alcohol research? processes in the cell. High-throughput technologies have What kinds of alcohol-related problems offered biologists tremendous opportu­ can we address using a systems biolo­ Q. MAX GUO, PH.D., is program direc­ nities but also have created considerable gy approach? What opportunities and tor and SAM ZAKHARI, PH.D., is director, challenges. How can we take advantage challenges are there in current and both at the Division of Metabolism and of this wealth of information to under­ future research? These are the kinds Health Effects, National Institute on stand its biological significance in health of questions that the articles in this Alcohol Abuse and Alcoholism, Bethesda, and disease? Systems biology is an special systems biology issue of Maryland.

Vol. 31, No. 1, 2008 5 for alcohol research, and discuss ways and tissues simultaneously. mere collection of components. opportunities and challenges facing Because the approaches taken by theory- Rather, they are derived and shaped alcohol researchers. oriented systems biologists are very by millions of years of evolutionary abstract and require specific expertise selection for survival, fitness, and in computation and modeling, early reproduction. For example, the What Is Systems Biology? systems biologists in the field of alcohol mitotic spindle5 in a cell is not just research will most likely adopt the created from simple polymerization Systems biology is a new scientific pragmatic approach. of α- and β-tubulins.6 It involves the discipline that studies the behaviors interaction of the polymerized micro­ of complex biological organizations tubules with motor proteins and the or processes through the integration What Is a Biological communication with many other cel­ of diverse quantitative information System? lular signals to accurately segregate and mathematical modeling to gener­ the sister chromosomes. The emer­ ate a predictive hypothesis on the “Biological system” is a loosely defined gent properties during chromosome functions of the dynamic biological term. To understand what constitutes segregation cannot be achieved by system (Aderem 2005; Auffray et al. a biological system, it is helpful to the collective properties of tubulins, 2003; Hood et al. 2004; Kirschner revisit a visionary paper by Hartwell microtubules, motors, or chromo­ 2005; Liu 2005; O’Malley and Dupre and colleagues (1999) published before somes alone. The spindle can be con­ 2005; Weston and Hood 2004). systems biology became a household sidered a functional module, which Systems biology may have quite term for biologists. Although the communicates and interacts with and different meanings to different people. authors did not mention the phrase organizes many other molecules and In general, systems biologists can be “systems biology” in that paper, their structures. The amazing harmony of organized into two camps. In the first proposed concept of functional mod­ spindle behaviors during cell division camp, the “systems-theoretic biologists” ules provides the building blocks for is achieved through evolutionary think that the focus of systems biology defining a biological system. Hartwell is to elucidate system principles and and colleagues proposed that the func­ 4 An emergent property is the new characteristic gener­ properties of operation based on com­ tions of a cell are carried out by mod­ ated through the interaction of individual components. ponent interactions in the biological ules made of many different kinds of 5 The mitotic spindle is an assembly of microtubules, system (Bork and Serrano 2005; interacting molecules. They argued structural components in the cell, created during the O’Malley and Dupre 2005). To that these functional modules provide process of cell division known as mitosis. them, systems biology is very abstract a critical level of biological organiza­ 6 α- and β-tubulins are the proteins that make up micro­ and precise. The mere integration of tion. Functional modules are not a tubules. constituents in the system for under­ standing the emergent properties4 of component interactions is insufficient for these theory-oriented systems 600 biologists. However, the majority of today’s systems biologists, who can be 500 described as “pragmatic systems biol­ ogists,” are gathered in the other 400 camp (O’Malley and Dupre 2005). They use systems biology as a practi­ 300 cal but vague term, denoting the study of interconnected phenomena 200 as systems and the collective analysis of large amounts of diverse data by integration and modeling. Despite 100 Number of Publications Number of Publications these differences, the pragmatic and theoretic systems biologists share 0 some common ground. For example, 2000 2002 2004 2006 both agree that systems biology Year involves data integration and mathe­ matical modeling and that models Figure 1 Number of publications with “systems biology” as a key word in the generated through systems biology search of the ISI Web of Science. The data plotted are from 2000 to studies should be predictive. 2006. The Web address for the ISI Web of Science is: http://portal. Alcohol-induced disorders are very isiknowledge.com/portal.cgi complex, involving numerous path­

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Systems Biology and Alcohol Research selection, as evidenced by comparing phate [ATP] synthase) have 13 and generate a model of how Complex I the spindle formations during closed 16 subunits, respectively. Each of functions. Based on this model, we mitosis and open mitosis. During closed these complexes can be considered a should be able to hypothesize and pre­ mitosis in yeast, the spindle is formed functional module, consisting of vari­ dict how the function of the complex within the nuclear membrane, which ous interacting subunits. But each of would change if we perturb a specific does not break down during mitosis. these complexes also can be consid­ component in the system, say by However, in higher organisms, the ered a biological system if our goal is ethanol. For example, if the amount nuclear membrane breaks down dur­ to understand how each of the com­ of one nuclear-encoded subunit is ing mitosis and the spindle is formed plexes works. However, if our goal is rate limiting for assembly of the com­ without constrains of the nuclear to understand how ATP is generated plex, one may predict that the expres­ membrane (open mitosis). The func­ through oxidative phosphorylation,8 sion of this nuclear gene is tightly tion of the nuclear membrane for we would consider the entire electron regulated. Aberrant regulation of its spindle formation in yeast may have transport chain as a biological system. gene expression (e.g., by a mutation evolved into that of a new structure of a transcriptional inhibitor) could called the spindle matrix in higher cause an increased amount of the sub­ organisms (Tsai et al. 2006; Zheng A biological system unit and thus increased activity of and Tsai 2006). NADH dehydrogenase (Complex I) A functional module always pos­ consists of either a single in the mitochondria. When experi­ sesses emergent properties derived mentally tested, this prediction could from the interactions of individual functional module or prove incorrect, because the overex­ components. These discrete proper­ many interconnected pressed subunit also could be regulat­ ties could not be predicted by study­ ed through degradation or transport ing the individual components in iso­ modules that work in into mitochondria. Based on the new lation. Although a functional module experimental data, the model would is a distinct entity separable from the concert to carry out the be further refined and improved. This functions of other modules, it is not functions of a biological integration of experimentation, data necessarily a rigid and fixed structure. processing, and mathematical modeling A component in one module also organization or is an iterative process, which continues could belong to another module. In process. until a relatively detailed and suffi­ addition to being insulated from each ciently accurate model is achieved. other in order to carry out many Despite different levels of complex­ diverse functions, functional modules In this case, the five complexes would ity in various biological systems, they are interconnected. The interactions be considered interconnected modules all share some common characteristics of these interconnected modules in in the system. With the same logic, we (Aderem 2005; Westerhoff and Palsson turn generate a higher layer of func­ can consider the mitochondrion, the cell, 2004). First, all biological systems tions at another biological level. organs, and organisms as larger bio­ display some forms of modularity. Recognizing functional modules logical systems with increasing com­ These modules are interconnected, as the building blocks of a biological plexity. The functional modules are often in a hierarchical structure. Second, organization or process, we could define interconnected in a hierarchical struc­ a biological system always possesses a biological system with the terms ture to form subsystems and systems. some emergent properties. The study used in modular biology. A biological If we use the systems biology of any participating component alone system consists of either a single func­ approach to study how Complex I would not be able to predict these tional module or many interconnected functions, we would gather information new properties. For example, studies modules that work in concert to carry on the identity of each component; of a subunit in mitochondrial Complex out the functions of a biological orga­ the regulation of its gene expression; I can never predict how the complex is nization or process. To illustrate this its synthesis, degradation, and trans­ assembled and how NADH dehydro­ point, we will use mitochondria as an port; and its interaction with other genase functions. Similarly, studies of example. The mitochondrial electron subunits, membranes, and molecules, Complex I alone can never predict how transport chain7 consists of five enzyme etc. In addition, we also would need ATP is generated through the electron complexes. Complex I (NADH­ to collect cellular, physiological, and transport chain. Finally, biological coenzyme Q reductase) contains clinical measurements associated with 45 subunits; Complex II (succinate­ the perturbation of the complex. We 7 The mitochondrial electron transport chain is composed of enzymes (Complexes I, II, III, and IV) that transfer coenzyme Q reductase) contains 4 also could collect data on the struc­ electrons and produce a proton gradient, which drives the subunits; Complex III (coenzyme tural and behavioral alterations of the synthesis of ATP by Complex V. Q-cytochrome c reductase) has 11 complex in response to external cues. 8 Oxidative phosphorylation is a metabolic pathway that subunits; Complexes IV (cytochrome These data then could be integrated uses energy released by the oxidation of nutrients to c oxidase) and V (adenosine triphos­ and computed mathematically to produce ATP.

Vol. 31, No. 1, 2008 7

systems are robust. A biological sys­ is indeed a systems biology disorder. study the quantity of RNAs and their tem is not merely a collection of its Alcohol research using a systems biology alternative splicing. In the past several parts. As described in the above approach will prove fruitful in unrav­ years, a significant amount of microar­ example, the mitotic spindle, as a bio­ eling the mechanisms of complex ray data has been generated and is logical system, is not formed through alcohol-induced disorders. reaching the critical mass that is simple polymerization of tubulins. A required for systems biology studies. dysfunctional spindle would cause At the protein level (i.e., pro­ detrimental consequences for the cell, Components of Systems teome), proteomic technologies can such as cancer or apoptosis. A biologi­ Biology for Alcohol systematically study the identity, cal system is a survivor of furious Research quantity, modification, localization, competition and natural selection Genomics and functional genomics are interaction, and function of all pro­ over millions of years. Over time, the driving forces behind the systems teins in a cell, often in a high- organisms have evolved to develop biology approach for alcohol-induced throughput manner. The proteomic many complex mechanisms, such as disorders, as discussed in other articles approaches would extend the power positive and negative feedback, to in this issue (see figure 2). At the DNA of the global gene expression analyses maintain the functional stability of level (genome and epigenome), vari­ for the following four primary rea­ their biological systems. Therefore, it ous global genomic technologies and sons: (1) RNA expression sometimes is no surprise that our biological stud­ does not correlate with protein expres­ ies often amaze us with the robustness sion, which is a more critical indica­ and harmony of biological systems. tor of the gene activity; (2) the activi­ At the systems ty of a protein could be significantly modulated by post-translational mod­ Alcohol-Induced level, alcohol affects ification, which cannot be reflected at Disorders Are Systems a variety of organs, the RNA level; (3) the function of a Biology Diseases gene is executed by its protein product biochemical or in a specific subcellular compartment; Alcohol’s actions in our body are very and (4) a protein molecule provides complex, and alcohol-induced physi­ signaling pathways, many more targets for regulation ological or behavioral changes are and other biological (quantity, functional activity, struc­ affected by a variety of intrinsic and ture, posttranslational modification, environmental factors. Alcohol-induced processes. localization, etc.), whereas RNAs most­ disorders, including organ damage ly are targeted for their quantity. In and addiction, reflect the genetic and 9 addition, proteins normally are more epigenetic makeup of an individual information can be applied to study stable than RNA in both cells and and the cumulative responses to alcohol genetic variation, gene mutation, gene body fluids. The stability and multi­ exposure and environmental pertur­ mapping, and genetic or epigenetic ple layers of regulation of proteins bations over time. Each of these factors regulation. Both genetic and epige­ make them much better candidates as may contribute only a small fraction netic mechanisms are crucially biomarkers or targets for therapeutic to the symptoms or phenotypes. At responsible for susceptibility, initiation, interventions. the molecular level, the effects of progression, and pathogenesis of At the metabolite level (i.e., meta­ alcohol and its metabolites are the alcohol-induced disorders. Evidently, bolome), metabolomics involves a consequences of changes in DNA, approaches to the study of gene regu­ detailed quantitative analysis of RNA, proteins, metabolites, and other lation purely based on gene sequence low–molecular weight metabolites molecules. At the systems level, alcohol would not be sufficient to explain over changing environmental condi­ affects a variety of organs, biochemical alcohol-induced pathogenesis. A better tions (e.g., alcohol administration) in or signaling pathways, and other bio­ understanding of epigenetic mecha­ a biological system. Metabolites are logical processes. This high degree of nisms will complement information the intermediate and end products of complexity in alcohol-induced disorders obtained from genetic, genomic, and cellular functions, and their levels and renders the traditional gene-by-gene functional genomic studies. The modulation are definitive reflections or single-discipline studies limited information can be integrated with other of an organism’s response to genetic because they only provide a fragmented experimental and clinical measurements or environmental perturbations. The view of a very complex picture. An to identify complex systems-level determination of these metabolites integrated approach, such as systems responses to alcohol and environmental biology, is essential in revealing a glob­ perturbations. 9 Epigenetics is the study of heritable changes in gene al picture of the perturbations leading At the RNA level (i.e., transcriptome), function that occur either to DNA without altering its sequences or to DNA-associated histone proteins. These to disease. From either pathogenetic DNA microarray and other global epigenetic changes can either be inherited mitotically from or etiological perspectives, alcoholism genomic technologies can be used to cell to cell or meiotically from generation to generation.

8 Alcohol Research & Health Systems Biology and Alcohol Research can be achieved by using a variety weight metabolites, compared with genomic and functional genomic data of tools such as mass spectrometry, more than 6,000 genes. and to extract their biological and nuclear magnetic resonance spec­ Another emerging area for systems clinical significance. The integration troscopy, capillary electrophoresis, and biology is glycomics, which is a global and modeling of diverse information, high-performance liquid chromatog­ approach to study complex carbohy­ including other biological and clinical raphy, in conjunction with a wide drates (or glycans) for their structure measurements, would vastly enhance range of bioinformatic, statistical, and function and their interaction the power of any single-discipline and computational tools. A virtual with other carbohydrates, proteins, approach, help to decipher the mech­ snapshot image can be obtained of lipids, and nucleic acids. Carbohydrates anisms of alcohol-induced disorders, the myriad of small molecules within and their interaction with other and provide new avenues for their the biological system and how these molecules are involved in a wide spec­ prognosis, diagnosis, and treatments. molecules are modulated in individu­ trum of cellular functions. Glycomic al time frames. Metabolomic studies studies may reveal glycan changes in alcohol research are complemen­ and provide novel avenues for under­ Challenges and tary to studies of the genome, tran­ standing alcohol’s actions, especially Opportunities scriptome, or proteome, because they on posttranslational modifications It is clear from the articles in this issue can extract latent biochemical infor­ of proteins. that well-developed systems biology mation of diagnostic or prognostic For systems biology studies, bioin­ programs are needed in the alcohol value, reflecting actual biological formatics, computation, statistical field. This is true in many other fields events, and can serve as a sentinel for analysis, and mathematical modeling as well, such as pulmonary diseases, diseases. Metabolomics also offers are all pivotal for integrating and diabetes, and aging. Interestingly, of approachable solutions because there making sense of large and complex the 575 systems biology papers pub­ are far fewer metabolites than genes datasets generated through the high- lished in 2006, many were reviews or gene products (RNAs and pro­ throughput -omic technologies (see and meeting abstracts. Currently, com­ teins) in many organisms studied. figure 2). Through integration and prehensive systems biology studies For example, in yeast, there are only modeling, these studies would allow still are quite uncommon in most approximately 600 low–molecular us to better exploit the complexity of biological and biomedical fields. Ob­ viously, systems biology approaches are difficult and present a variety of new challenges. This section focuses Genetic Biological Epidemiol. Clinical on a few of these challenges that are Studies Studies Studies Studies relevant to alcohol research. First, alcohol research still is facing

Alcohol Alcohol- many technological challenges. As Animal Human Output Induced & Input Models Subjects discussed above, genomic and func­ Environment Disorders tional genomic studies are the driving forces behind systems biology, which Genome Mechanism Epigenome Transcriptome Prognosis demands the availability of sufficient Diagnosis and reliable data from many different Treatment Proteome Metabolome technological platforms (also see Ge et al. 2003). Not only do we need to Glycome expand our repertoire of technologies to include epigenomic, metabolomic, glycomic, and other –omic approach­ es, we must also integrate these tech­ nologies in systems biology studies. Second, as with all other fields of Figure 2 Systems biology approach for alcohol research. The diagram empha­ biomedical research, the bioinformat­ sizes the concept of integration and the significance of bioinformatics, ics, computation, and statistical or computation, and mathematical modeling. The lines between the “omes” are meant to indicate that the web of data on all the “omes” should be mathematical modeling necessary for integrated and collectively analyzed. Genomic and functional genomic systems biology studies present even data also should be integrated with genetic, other biological, epidemio­ greater challenges. Finally, additional logical, and clinical studies. The ultimate goal of systems biology studies technological obstacles, such as stan­ is to understand the mechanisms and develop better prognosis, diagno­ dardization; creation of uniform, sys­ sis, and treatment for alcohol-induced disorders. tematic vocabularies and systems of annotation; and digitalized output of

Vol. 31, No. 1, 2008 9 other biological and clinical measure­ investigators on a single grant, repre­ References ments need to be addressed (also see sents a move in the right direction. Cassman 2005). Because these chal­ Finally, systems biology studies often ADEREM, A. Systems biology: Its practice and chal­ lenges are not specific to individual are hampered by the lack of trained lenges. Cell 121:511–513, 2005. PMID: 15907465 investigators, they should be tackled investigators with interdisciplinary AUFFRAY, C.; IMBEAUD, S.; ROUX-ROUQUIE, M.; through collaborative and concerted talents and skills. A fascinating aspect AND HOOD, L. From functional genomics to systems efforts. of systems biology is that it integrates biology: Concepts and practices. Comptes Rendus In addition to these technological large amounts of diverse forms of Biologies 326:879–892, 2003. PMID: 14744094 challenges, we need to address some information for mathematical model­ BORK, P., AND SERRANO, L. Towards cellular systems unprecedented organizational chal­ ing. Therefore, systems biology neces­ in 4D. Cell 121:507–509, 2005. PMID: 15907464 lenges. Traditionally, biomedical sitates the involvement of computer CASSMAN, M. Barriers to progress in systems biol­ research is carried out in individual scientists, statisticians, and mathe­ ogy. Nature 438:1079, 2005. PMID: 16371982 laboratories with relatively focused maticians who also are well-versed in GE, H.; WALHOUT, A.J.; AND VIDAL, M. Integrating areas of inquiry. When collaborations biology, as well as biologists who can “omic” information: A bridge between genomics are involved, they are normally simple understand bioinformatics, statistics, and systems biology. Trends in Genetics 19:551– 560, 2003. PMID: 14550629 and limited to a few experiments. and mathematical modeling. The bot­ Even for a very large endeavor such tleneck of the systems biology approach HARTWELL, L.H.; HOPFIELD, J.J.; LEIBLER, S.; AND as the Human Genome Project—in is the cross-disciplinary training for MURRAY, A.W. From molecular to modular cell which collaborations were extensive— biology. Nature 402 (6761 Suppl.):C47–C52, a new generation of researchers who 1999. PMID: 10591225 organizational structure was clear and can tackle the multidisciplinary com­ management was relatively straight­ plexity with ease. Universities and HOOD, L.; HEATH, J.R.; PHELPS, M.E.; AND LIN, forward. For example, each chromo­ B. Systems biology and new technologies enable research institutions need to create some was sequenced by an individual predictive and preventative medicine. Science multidisciplinary training programs. 306:640–643, 2004. PMID: 15499008 sequencing center and all sequence This issue also needs to be addressed information was easily combined into KIRSCHNER, M.W. The meaning of systems biol­ a single database. Systems biology has by funding agencies. ogy. Cell 121:503–504, 2005. PMID: 15907462 Despite these challenges, compre­ to deal with much more diverse for­ LIU, E.T. Systems biology, integrative biology, pre­ mats of data, many more different hensive systems biology is a realistic dictive biology. Cell 121:505–506, 2005. PMID: technologies, and much greater com­ prospect now for the studies of alcohol- 15907463 induced disorders. Systems biology, with plexity in terms of both biological O’MALLEY, M.A., AND DUPRE, J. Fundamental system and organizational structure. the help of continuing technological issues in systems biology. BioEssays 27:1270–1276, Experts involved in collaborative sys­ innovations and multidisciplinary 2005. PMID: 16299766 tems biology studies most likely teamwork, provides the integrative TSAI, M.Y.; WANG, S.; HEIDINGER, J.M.; ET AL.A reside in different departments, if not approach necessary for the future mitotic lamin B matrix induced by RanGTP prognosis, diagnosis, and treatment required for spindle assembly. Science 311:1887– different institutions or companies. 1893, 2006. PMID: 16543417 Investigators need to overcome many of alcohol-induced disorders. ■ intrinsic and extrinsic barriers to WESTERHOFF, H.V., AND PALSSON, B.O. The evo­ form a multidisciplinary team aimed lution of molecular biology into systems biology. Acknowledgments Nature Biotechnology 22:1249–1252, 2004. PMID: at understanding the function of a 15470464 biological system. Research institu­ We thank Dr. Ting-Kai Li for his helpful WESTON, A.D., AND HOOD, L. Systems biology, tions and funding agencies also need comments and suggestions. proteomics, and the future of health care: Toward to take the necessary steps to encour­ predictive, preventative, and personalized medicine. age and facilitate the sharing and team Journal of Proteome Research 3:179–196, 2004. work that are essential for a systems Financial Disclosure PMID: 15113093 biology approach. A recent change of ZHENG, Y., AND TSAI, M.Y. The mitotic spindle policy by the National Institutes of The authors declare that they have no matrix: A fibro-membranous lamin connection. Health, which allows multiple principal competing financial interests. Cell Cycle 5:2345–2347, 2006. PMID: 17102624

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