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Cellular Participation In Physiology Jim Pierce Bi 145a Lecture 3, 2009-2010 Cellular Physiology There is more to cells than just performing basal functions. Cellular Physiology is the study of how cells perform both basal and tissue specific functions. It is the bridge between molecular biology and tissue function. Cellular Physiology General Cellular Physiology The study of inputs, metabolism, and outputs. Our goal for this lecture is to look at examples of inputs and outputs. Cellular Physiology Specific Cellular Physiology This is the study of specific aspects of cellular function. Our goal in this lecture is to discuss some of the common functions: membrane function, cellular structure, cellular motors, and secretion. General Cell Physiology Homeostasis versus Active Function Often, we want to do more than “cruise along” at status quo. This means doing stuff Homeostasis What is homeostasis? It is central to understanding physiology, and makes metabolism much easier. It is the property of a system to try to maintain constancy in the face of external perturbations. Homeostasis Regulation When a system maintains some variable despite internal or external perturbation. Control When a force adjusts the output of a system over time. Refrigerator Thermostat Glycogen in a Myocyte Homeostasis Figuring out the Refrigerator is easy The thermostat controls The “dial” indicates the set point A circuit in the Refrigerator regulates. It compares the actual temperature to the thermostat’s set point It heats or cools accordingly Refrigerator Cooling Homeostasis Glycogen is not so easy. What controls glycogen levels? There is no thermostat. The myocyte does not receive a specific “glycogen level signal.” Homeostasis What controls glycogen levels? There are external signals that inform the myocyte of the state of the body. There are internal signals that inform the myocyte of the state of the cell. The cell must integrate these signals to make that decision. Homeostasis The way a cell integrates these signals to make that decision is called A Control Structure Homeostasis How is glycogen level regulated? Because there is no thermostat, it is not a simple “compare” like the fridge Instead the pathway “pays attention” to every reaction, intermediate, and final product This is called a Regulatory Structure Homeostasis How is glycogen level regulated? The Regulatory Structure is affected by the Control Structure. It is through those interactions that the Regulatory Structure obeys the Control Structure Homeostasis What makes up these structures? Cells Do Chemistry. (they do physics, too, but I don’t like physics so I won’t talk about it) Chemical Compounds (stuff) Processes Involving Chemical Compounds (a way to change stuff) Homeostasis Structures are composed of reactions AB CD EF Metabolism Xsource supply demand --> M --> (in a nutshell) Xsink Regulatory Network Xsource supply demand --> M --> (in a nutshell) Xsink Control Network Xsource supply demand --> M --> (in a nutshell) Xsink Why is this confusing? The final product has the greatest effect on the flux through metabolism So the USE of the final product exerts CONTROL Regulation versus Control So certain Regulatory structures give “Control” to the end product This is probably why biologists use “regulate” and “control” interchangeably Just remember, like precision and accuracy, control and regulation are different! Metabolism and its Control So how do we describe these things? We start with a Model of the system Metabolic Pathway A B C D A set of metabolites and reactions involving those metabolites. (note that a metabolic pathway can be described by graph theory) Generalized Linear Metabolic Pathway A B C D Generalized Branched Metabolic Pathway Generalized Substrate Cycle Metabolic Network Control and Regulation We can then describe how any given thing (enzyme, molecule, extrinsic parameter) affects any other given thing This gives a bunch of variables This allows a mathematical model Control in Dynamical Systems Models That Exist: Linear Systems Linear Models Non-Linear Systems Non-Linear Models Linear Models Metabolism There is more to metabolism than just the graph of the reactions Location of the reactions Cytosolic, Membrane Bound, Nuclear Job in “the bigger picture” Anabolism versus Catabolism Control by Supply Feedforward Control: 1) Can achieve high control of flux 2) High control of flux forces us to have low control of metabolites! (That means AMPLIFICATION) Control By Demand Feedback Control 1) Can achieve high control of flux 2) High control of flux forces us to have high control of metabolites! Metabolic Control Analysis We can prove that: 1) Feedback is the way we get control of both flux and concentration. 2) Feedforward is the way we get control of flux and amplification. Internal State The first consideration in “doing stuff” is the Internal State of the Cell The set of DNA, RNA, and protein (especially “transcription factors”) Organelles and Compartmentalization Membrane and its Potential Secondary messengers Internal State Sometimes, the decision to Activate is the Internal State The best studied example is Cell Cycle There is an internal clock (multiple, actually) In many cases, the ticking of the clock alone is the largest stimulus for cell division Internal State A closely related (and more interesting) example is Early Development Much of the earliest patterning results from internal state Distribution of Bicoid mRNA (Drosophila) Distribution of Vg-1 protein (Xenopus) Random Genetic and Positional Noise (Chick rotates with gravity, Mouse random based on position in ICM) (Bi 182 for more info!) Internal State Other basal functions include: Basal secretion in glands Basal membrane potential patterns Anabolism and Catabolism for Housekeeping Internal State In the case of cell cycle, the output includes: Replicating DNA and organelles Nuclear Division Cytosol Division A huge number of checkpoints Lots of error correcting Internal State In the case of early development the output consists of: Spacial and Temporal Patterning of space (intra and extracellular) Interpreting “Internal state” into Cellular Phenotype General Cell Physiology Obviously, though, internal state cannot be the only cue! In a complex organism, even making ATP depends on the state of the organism! (a fat cell should never steal glucose from a starving brain cell) General Cell Physiology Types of Inputs: Small Molecules Neurotransmitters, Steroids, Peptides Non steroid, non peptide hormones General Cell Physiology Types of Inputs: Large Molecules ICAMs, Selectins, Integrins Lipoproteins Other Immunoglobulins Other Glycoproteins Small Molecules Why Small Molecules? They are very versatile They can carry information (in both concentration and concentration gradient) They can diffuse or be transported. Small Molecules Why Small Molecules? They are very efficient The earliest “computation” on small molecules was probably bacterial chemotaxis Food (i.e. reduced molecules) was transduced into swimming behavior Small Molecules Why Small Molecules? They are very efficient This system can be harnessed by using specific small molecules as a signal Ever notice that many neurotransmitters are decarboxylated amino acids? Small Molecules Examples of Small Molecules Addition / Integration Two inhibitory cells both release GABA onto the same dendrite, increasing hyperpolarization Each parathyroid cell releases hormone into the blood, and response is a function of “total hormone” levels. Small Molecules Retention Insulin binds to its receptor and is internalized, providing continued signaling. Degredation Serum Catecholamine-O-Methyl-Transferase has different rates of catecholamine removal than neuronal reuptake machinery Small Molecules Gradient Retinoic Acid (vitamin A) and HOX genes DPP in certain non-mammal animals Target Autocrine – stimulates self Paracrine – stimulates neighbor Neurocrine – neural synapse Endocrine – stimulates distant cell via blood Neuroendocrine – neural secretion into blood Large Molecules Why Large Molecules? They can “mark” an area of extracellular space. (i.e. they stay put) They convey information about tissue structure (both cell-cell and cell-ECM). Large Molecules Consider Neural Crest Cells Early development “encodes” space with a set of small molecules, gradients, and large molecules. Neural crest cells migrate through this space, using the cellular computer to respond to spacial differences Large Molecules Consider a skin injury… The cells at the edge of the injury lose the suppressing signal from cell-cell adhesion receptors. …But they cannot grow without the stimulating signal from the basement membrane. Large Molecules Thus, these large signals are key to tissue functioning. We spend so much time thinking about the small signals (Bi/CNS 150) that we sometimes forget how much information is encoded in these large molecules. Large Molecules Every time two cells stick together, they are communicating Every time a cell sits in the extracellular matrix, it is listening to its surroundings General Cell Physiology Also remember: a huge portion of the signals are suppressive. In the brain… On the basement membrane… In the glands... General Cell Physiology Secondary Messengers yuk. Secondary Messengers Words of advice about these guys: Anything can function as a secondary messenger if it can convey information, such as small molecules, assembled structures, and even the membrane itself. Secondary Messengers Always think about the cellular compartment where the secondary messenger is located; different compartments have different properties. The surface of the membrane is two dimensional, and therefore is better for diffusion. Cytosolic messengers can overcome three dimension diffusion by assembly. On the flip side, cytosolic messengers can also change compartments and locations in the cell. Secondary Messengers Do not fall into the trap of thinking of certain messengers as “activating” or “suppressing.” cAMP cGMP Ca++ Secondary Messengers “Secondary messengers” only get their name because they're supposedly restricted to the cell itself. Some hormones (steroids) compute like secondary messengers Some secondary messengers (nitric oxide) can change cells like hormones. General Cell Physiology Concept Questions? Cancer We talked about: Homeostasis Regulation versus Control How one could actually study it Now: Cancer Cancer Tumor Tumere – “swelling” What can cause swelling? Too Many Cells Too Much Extracellular Matrix Too Much Fluid Neoplasm Neo – “new” Plasm – “growth” Too Many Cells Cancer Neoplasm Benign – Cells stay where they are Malignant – Cells invade somewhere new Often benign ends in –oma (lipoma) Often malignant ends in -carcinoma = malignant from epithelium -sarcoma = malignant from meso/endothelium Cancer Is benign “benign?” A benign fatty growth that squishes the trachea and suffocates the patient Is malignant “malignant?” A slow growing skin cancer that never causes any symptoms and the patient dies 20 years later of a heart attack Cancer Cancer Multi-Hit Theory Cancer doesn’t develop overnight After “watching” many different tumors, one begins to notice a progression. Over time, the tumor gets uglier, bigger, grows faster, and grows in new places. Multi Hit Theory The multi-hit theory was proposed simply by watching the DNA The older or more severe the tumor… … the more DNA mutations that could be found. Multi Hit Theory So it was hypothesized that cancer develops by sequential epigenetic mutations In that case, a predisposition to cancer occurs from germ line mutations, which is how many important genes were found Multi Hit Theory Further Support arrived with the identification of viruses that induce cancer These viruses contained mutated genes v-myc (v-genes in general) = Viral myc c-myc (c-genes) = Cellular myc Multi Hit Theory These viral genes were called Oncogenes – (onko (greek) – mass ) Their corresponding cellular genes were called “proto-oncogenes” (proto – first) They behave (generally) as gain-of-function phenotype Multi Hit Theory Another class of genes were described that provided “resistance” to carcinogenesis from viruses These genes were called Tumor Suppressor Factors They behaved (generally) like loss of function phenotype Multi Hit Theory So the multi-hit theory is the idea that cancer arises through a series of steps Each one corresponds to a “gain of function” or “loss of function” mutation in a specific gene Thus explaining what surgeons had been observing since Brahman period medicine in early India Multi Hit Theory But that implies that cancer is “Growth out of control” Uncontrolled Cell Cycle, which accelerates and accelerates. Cancer So why is it so difficult to grow cancer cells in a dish? Cancer Primary Cultures Died if they weren’t attached to a surface Died if there were too many Died if there were too few Died without serum or growth factors Died with too much serum or growth factors …And often just died anyway Cancer HeLa cells 1951 – Johns Hopkins Medical School Henrietta Lacks, mother of four Cervical Cancer cells were cultured by George Gay, MD without permission. They grew “horrifically” Cancer Prior to HeLa cells, primary cultures of human cells had a “finite” lifespan Just to keep them alive for a week took the addition of “serum” with its panoply of unknown factors. Cancer So how could cancer be so awful, if it won’t even grown in a dish? The answer lies in “phenotypes” Cancer is better thought of as a “Disease of Gain-of-Phenotype” Cancer There are many different phenotypes: Needs basement membrane neighboring cells growth factors Abilities Secretion, Absorption Metabolism Cell Cycle Computation Cancer These include “new” or “unusual” phenotypes: Invasion through basement membrane Ability to migrate Ability to live in a new milieu Blood Lymph Nodes Other Tissues Cancer Evasion of Immune System Resistance to killing (similar to viruses) Resistance to Aging Telomeres DNA Damage Cancer Secretion of Hormones Autocrine Stimulation Growth Factors Recruitment of “support” (angiogenesis) Cancer So every tumor is a different combination of “phenotypes” Disease progression is sequential addition of new phenotypes, each which result from mutations Cancer External / Internal signals for survival Gaining proliferative phenotype Losing apoptotic / senescent phenotype Gaining metastatic phenotypes Avoiding immune respone Angiogenesis and other novel phenotypes Cancer These mutations often make the cell appear less differentiated and more multipotent. So in many ways, understanding Cancer is very similar to understanding Stem Cells (and their differentiation phenotypes) Cancer For more on Clinical Aspects of Cancer Try out my brand new Bi 23, Winter 09-10 Cancer Examples Esophageal Adenocarcinoma Fastest Rising Western Cancer (~500% in the last 30 years) Normal Esophageal Mucosa Normal Layers Esophageal Cancer Chronic gastroesophageal reflux Leads to Acid and Bile Exposure The cells will try to protect themselves Barrett’s Esophagus With Esophagitis Esophageal Cancer To defend itself, the cell “gains” the phenotype of “mucous secretion” Normal Barrett’s Esophageal Cancer This is called “metaplasia” - Metaplasia is when one tissue type changes to another tissue type that “naturally” occurs in the body in a different location Metaplasia requires that the cell respond to stimuli and change phenotype Esophageal Cancer The mechanisms of metaplasia depend on the tissue and stimulus, but often involve DNA damage In the case of Esophageal Cancer, bile acids are carcinogenic Skin Cancer – UV light Lung Cancer – Smoking Cervical Cancer – HPV Esophageal Cancer Not surprisingly, continued acid exposure allows progressive accumulation of DNA damage It becomes carcinoma when: It grows without regard to neighbors It is able to cross the basement membrane Esophageal Cancer Barrett’s Adenocarcinoma Adenocarcinoma Cancer Progression What we see is: Primary Injury (bile) Progressive DNA damage Gain of Phenotypes “Pre-Cancer” – not yet across the Basement Membrane “Cancer” – crossed over Metastasis – gone somewhere else Good Cancer / Bad Cancer Why focus on cell cycle and apoptosis? Removing checkpoints and error correcting facilitates gain-of-phenotype Apoptosis is the solution for excessive DNA damage, broken apoptosis leads to proliferation despite severe damage. Questions?