Quiz 1 due Monday Aug29th 11:59pm   (covering safety/microscopy/sterile technique AND reading for week of this week)
Unit 1: Learning techniques and Determining our baseline of response (Defining 'normal')
Unit 1: 'The Signal 'and How Yeast Respond to It :
Techniques for characterizing yeast mating reaction 

Quiz 4 due Monday Sept 19th by 11:59pm
Unit 1: 'The Effect' Part III Differential gene expression Detecting and quantitating gene expression patterns .
Work on Materials and Methods.
DUE Friday 9/23 before noon: Material and Methods

Mini unit: How do scientists communicate? Grants and Funding

Unit 2: Applying techniques to novel condition/ Comparing the 'expected' to the 'unknown'
Characterizing the mating abnormalities in two strains of yeast.
Groups meet to plan study of mutants/ Written outline of approach must be approved before leaving lab 

ALL Lab Articles due Wednesday 11/30 Before 10am
Lab cleanup, Course evaluations

Introduction to course, Overview of case studies
READING (yes, for today)
The Practice of Science At the Edge of Knowledge 
By Frederick Grinnell  http://www.faseb.org/opar/newsletter/6x00/guest.htm
What is the role of methodology in science? of intuition? of luck? of good PR?

Background: Why Cell biology is more than histology: Biological Membranes and Cellular components

Two classes reviewing background-- Need know the players before anything else.
In chapter 10 focus on the roles and components of biological membranes (pg583-602)
CBI animations and videos-- Use these to see the components of a membrane. The keratocyte is an example of membrane fluidity. Notice how it can re arrange the components to generate force and move.
The Virtual Cell (http://personal.tmlp.com/Jimr57/textbook/chapter3/chapter3.htm)
Make sure you are familiar with all of the cellular components and their functions. Explore the site. Note when you click on an organelle the popup window has navigation buttons at the top--use them to zoom in and out.
In Chapter 12 focus on how organelles are 'connected' (p659-666)

What role do biological membranes play in the cell? What are the components that make them up? How are they important to its function? What are the fluid mosiac model and lipid rafts? How do biological membranes define organelles and what the organelles? (Is everything in the cell an organelle? what kinds of cells have organelles and what kinds do not?)

Cellular components part II: No Cell is an island
Introduction to Case Study I: Grave's Disease-- Symptoms. Causes?

What are the functions of organelles and other cellular structures? Think about the interplay or connections there between a cell and its environment? Compare and contrast this interaction for a multicellular organism vs a single celled one.

Communication is the key.
With Graves' disease and many other disorders the symptoms can be summed up by quoting the movie classic "Cool Hand Luke"-- 'What we have here is a failure to communicate'. The problem is that there are many ways to mess up communication-- what has appened in Graves' disease to cause excess secretion of T3 and T4? In the next four classes we will discuss 'normal' extracellular signalling, how the mechanisms cells use to perceive those signals and what happens once the signals get in. During this section keep two questions in mind-- 1) How could these molecular mechanisms be altered to result in the symptoms seen in Graves' disease and 2) How does this relate to the mating interactions we are examining in lab?

I. Extracellular signaling and the receptors that mediate it.

What are the types of extracellular signaling? Which types might be faster/slower (why)? Given the diagram of normal thyroid signaling what types of extracellular signaling are being used? What are the types of cell surface receptors? (Some signaling molecules don't use cell surface receptors-- how do they get in?) Given the symptoms of Graves' disease what might be going wrong in the thyroid signaling pathway? (Think of more than one possibility and how you might test it)
In class we will discuss the exact types of receptors involved in pathway connected to T3 and T4 release.

II. So many signals--how does a cell know when to react? Specificity and the receptor

Interactions require the proper fit between receptors and ligands. Since all of the receptors we know are proteins we need to discuss proteins and protein folding before we can get to binding specificity.

What are the amino acids and how do they come together and fold up into proteins? What are the levels of protein structure? What ways can you think of to change a protein's folding? What are binding sites and active sites? How do antibodies (CBI 24.1) exemplify binding specificity? Knowing that TSH, T3 and T4 are the hormones involved and TSH receptor is found on the thyroid cells and is activated by TSH what structural changes could result in an overstimulated thyroid?

The ligands (signaling molecules) we have discussed are outside the cell. The plasma membrane is a barrier that these hormones cannot cross. In this class we will discuss how the receptor-ligand interaction allows a signal to cross the membrane even though the ligand stays 'outside'.

The Chapter 10 reading reviews membrane proteins like the TSH receptor. The enzyme reading and CBI animations show the effects of interactions between molecules.

What is the relationship between a proteins's conformation (shape) and its function? TSH is polar (can't get across the thyroid cell membrane by itself) how could it have an effect on the thyroid cell? Why don't all cells react to TSH? What could be happening to cause the thyroid to be overstimulated in Graves' patients?

IV. Response Cascades and 2nd messengers.

Activation by another means & Introduction to paper discussion (how we'll do it)

Think about the connection between antibody form and function. How does the body realize that it 'should' mount an immune attack? What happens during an immune response? What happens (to the body) when the immune system loses the ability to differentiate between self and non-self? How could autoimmune diseases be related to an overactive thyroid?

Introduction to paper discussion (how we'll do it)

Introduction to Case Study 2: Bipolar Disorder

Review parts of synapse/overview of normal nerve transmission
Background on Bipolar disorder will be presented. It is a mental illness that effects neuronal transmission resulting in wide variations in mood that are very disruptive to the individual's life.What is a neuron? A synapse? What are the types of synapses and how are signals passed across them? Think about the types of problems-- at the molecular level-- that could cause not just depressed or elevated moods but wide swinging of the pendulum between these two states.

 The players: Neurotransmitters and their receptors

Tale of 2 proteins: neuropeptide and neurotransmitter receptor
Protein #1: Cytosolic neuropeptide--How is it made?
(DNA/Transcription/Translation--in a day--should be a review)
If a person with bipolar disorder suffers from varied activity at a synapse then perhaps there is a problem with the way the signaling molecule is made or packaged. Neuropeptides and many neurotransmitters are made from amino acids. Neuropeptides are proteins and epinephrine and norepinephrine are tyrosine derivatives (so an enzyme--a protein-- modified the tyrosine). All neuronal signaling molecules must be able to diffuse across the synaptic cleft. So how do you make a diffusible protein? How do you get it into a synaptic vesicle so that it can be released during synaptic transmission?

Yes today's reading is long. Yes it covers alot of material but it should be review. DNA structure, transcription and translation are some of the topics you should have had at least twice in your biological career so we will move on to how these basic principles would relate to 'our' neurotransmitter at noradrenergic receptor. The animations are the short version-- focus on them and use the text to fill in rough spots.

What is the central dogma? How would a neuropeptide get made (in general terms)? What are the basic parts of DNA, RNA, and proteins? What is the difference between hnRNA, mRNA and tRNA? How does RNA polymerase work and what does it make? How does it know where to start and stop? How does a ribosome work and what does it make? How does it know where to start and stop? If the DNA in every cell in your body is the same why don't your adipose (fat) cells secrete epinephrine? If the DNA contains all of the information why doesn't the ribosome just 'read' it? Why have intermediate steps?

More protein manufacturing (finish translation)

Sorting the neurotransmitter-- some use direct access

T

R

T

Protein #2: Neurotransmitter receptor
How is it made? packaged? Intro to vesicles

Neurotransmitter receptors need to be part of a membrane-- they are integral membrane proteins (also called transmembrane proteins). Like other proteins the receptors are made in the cytosol. How do they get into the membrane? How can they be made in an aqueous environment and end up with parts in the hydrophobic membrane and parts on either side of that membrane? A simple 'door' through the membrane can't be enough. (A door to a room does not let you become part of the wall). The process of inserting membrane proteins is known as protein translocation and is part of protein sorting. How are the 'right' proteins chosen for translocation? (Said another way-- why is a neurotransmitter receptor synthesized in the cytosol and translocated but the parts of a trimer G protein are synthesized in the cytosol and remain there?) When does translocation take place? Does it occur during translation or after the protein has been made? Is there translocation machinery on every membrane or can newly made membrane proteins only get into a membrane at certain organelles? Moving a protein across a membrane is thermodynamically unfavorable (to say the least) where does the energy come from that powers translocation? Be able to describe the translocation mechanisms and think about how you would be able to tell which one was being used (what are their similarities and differences).

Didn't get to until
Thurs
10/27

How are the packages delivered? Secretion and the SNARE hypothesis

Now that the receptor is in the membrane and the neurotransmitter is in the synaptic vesicle how do we get them to the right place? Why do synaptic vesicles fuse with the plasma membrane of the nerve terminal and not with the plasma membrane at the nerve cell body? Why don't they fuse with the membrane of the lysosome? The 'easy' answer is that synaptic vesicles only fuse at the synaptic terminal because that's where the neurotransmitter is supposed to go. Releasing it anywhere else would be wasteful and possibly detrimental. The translocator gets the receptor 'into' the ER membrane but vesicles are needed to get the membrane protein to the dendrite membrane where it can function correctly. How do these vesicles form? How do they end up containing the right content? (things that are supposed to be delivered, not ones that should stay in the ER)? What is the difference in regulations level between constitutive and regulated secretion.

Other types of intracellular traffic.

Of course cells don't only exocytose they also endocytose, recycle and transcytose proteins. All of these forms of traffic use vesicles. How are the vesicles similar and different? Given any experimental method you wanted could you tell them apart? If no--why not? If yes--how?

How do the packages get there? Molecular roads and infrastructure.
Vesicles could just float around until they bump into the right membrane, but they don't. The speed of vesicular traffic testifies to the fact that brownian motion and diffusion aren't controlling the ways that vesicles move. Perhaps that's where mutations causing bipolar come into play--metaphorically speaking, maybe the parts are fine but the delivery trucks go on strike periodically or the roads are bad. What are the molecular road and delivery trucks? The cytoskeleton and molecular motors that move vesicles along the cytoskeleton.

Look at the video clip as an example of how the cell can change shape. Think about the kinds of structures that would be necessary to maintain those shapes. What are the components of the cytoskeleton? What do they have in common and how are they different. Are they rigid or flexible? Are they built once, remodelled sometimes or constantly changing? Why is this property important to cellular function? How could it effect cellular function if this property was altered? (Increased flexibility or increased rigidity)

(Paper #2 and questions are now available on Blackboard. Questions are due on Monday 11/7 BEFORE our discussion of the paper)

Molecular roads and infrastructure.
Regulation of cytoskeleton-- building and maintaining the roads and the engines that move vesicles
Reading

The cytoskeleton acts as both structural support and roads for the cell. The video clips illustrate the dynamic nature of the cytoskeleton. Also note how the heart muscle cell not only changes shape but is able to generate force through its shape changes. The leading edge of the migrating cell in 16.2 is extended by microtubules pushing it forward-- but the microtubules aren't stable. They grow and retract. How can that be? How can you have a constant structure if the skeleton keeps falling apart? How can vesicles end up in the right place if the roads break down? What are the processed of dynamic instability and treadmilling? How are they similar and different? Why is this instability important to the cell? Do cytoskeletal elements and the regulation of their stability seem like a good direction for study?

Now we have a little better understanding of the cellular 'roads'. Now we turn our attention to the engines that drive along those roads-- molecular motors. As with most cellular machines, molecular motors are made of proteins. These proteins have certain specificities and enzymatic activity that are important for their function. What are the motors? How do they 'know' what cargo to bind with and what roads to move along?
Do molecular motors seem like a plausible candidate for the cause of bipolar disorder? Why or why not?

Initial set of questions due Noon Monday 11/7
Discussion of Paper #2: C. McCann, et al. (2005) Peptide tags for labeling membrane proteins in live cells with multiple fluorophores. Biotechniques 38: advance publication no pages yet

Introduction to Case Study 3: Introduction to Werner's Syndrome

Life of a Cell: Cell cycle and its regulation

Final exam 'prequestions' will be emailed to you on Thursday

Telomere theory of Aging

Paper 3: Austriaco, Jr and Guarente 1997 PNAS vol94 9768-72
Beginning of discussion. Paper and Discussion questions available on Blackboard

Continue discussion of Paper 3: Austriaco, Jr and Guarente 1997 PNAS vol94 9768-72

Prefinal question list available on course website.

Why and how do cells die? Apoptosis and Death due to damage 

Both sections of the final exam will be emailed to you on Monday December 5th
The final is due Monday December 12th before 9am