PART-1

Comprehensive Exam- June 2012

Section-1 (Part A) 9AM 1PM
Instructions: Attempt 8 of the 15 questions in this section. You have to pass 6 (six).
Please spend no more than 30 minutes on each question. Begin the answer to each question on
a fresh page

Q1.
a. Adaptation is a fundamental property of signaling systems; e.g. when you enter a dark room, your
eyes eventually adjust to the low light level. Suggest three key molecular/cellular strategies (in any
biological context) by which a signaling system can adapt to time-dependent inputs.

b. For a signaling circuit capable of adaptation, plot the downstream output when the input has the
following form (you may assume that the adaptation mechanism operates on the timescale of several
minutes):

Q2.
A genome-wide screen in humans finds a recessive mutation in a gene with a strong link to
cardiovascular disease. The gene encodes a ubiquitously expressed protein with no predicted function.
Describe in some detail two different strategies that you could follow in your lab to establish the
function of this protein.

Q3.
a. In contrast to the case for soluble globular proteins, almost all residues in the transmembrane
portions of integral membrane proteins are ordered into secondary structure elements such as alpha
helices and beta sheets. Provide a biophysical explanation.

b. An alpha helix is a coiled structure with a pitch of 5.4 Angstroms, and 3.6 residues per turn. The
"hydrophobic thickness" of a lipid bilayer is about 30 Angstroms. In a few sentences, describe an
algorithm to identify putative trans-membrane alpha-helical segments of an integral membrane protein
of known sequence.

c. There are over 300 membrane proteins of known structure. Among these, it is observed that the
number of residues of trans-membrane alpha-helical segments can vary. What would you predict to be
the main structural difference between short and long trans-membrane segments?

Q4
a) Studies reveal that plants that live at high elevations tend to have a higher proportion of self-
fertilization. Can you think of reasons why this might be the case? What assumption would you be
making? (Hint: pollination)

b) Primula are high elevation plants that live in the Himalayas, among other places in the world. The
flowers only outcross, and do not self fertilize. However, the highest number of species co-occur at high
elevations. For example, elevations between 4000-4500 may have four species, while those at 3500-
4000 has two and only one occurs between 3000-3500m. How might multiple, non-selfing species co-
exist at high elevations?


Q5
A cylindrical non-myelinated axon is stimulated at both ends, so that action potentials propagate toward
each other. The axon has the usual HH channels Na and K delayed rectifier.

x
y






a) Draw the membrane potential Vm at the midpoint position x as a function of time
b) Draw Vm at y as a function of time, aligned on the time axis with the above graph.
c) Draw the open fraction of the inactivation gate h of sodium channels at y, as a function of time,
aligned with the above graphs.
d) Draw Vm vs. position, 1 msec after the APs have collided, that is, 1 msec after the peaks overlap.
e) Do the APs continue after the collision? With reference to these figures, explain.


Q6.
Dales principle holds that a single neuron uses one and the same substance as its transmitter across all
of its synapses. However, recent studies show that many neurons are capable of releasing more than
one transmitter (co-transmission), and sometimes the same neuron releases different transmitters from
different terminals. You are given a culture of medium spiny neurons from the striatum of the star-
nosed mole.
a) Describe at least two different methods by which you will determine the number of transmitters
that MSNs use.
b) How will you determine which transmitters are released at each contact? (Hint: MSNs are
known to contain GABA, enkephalin, dynorphin, and SubstanceP).


Q 7.
Tic-tac-toe, or Noughts-and-crosses, is a two-player game you are probably familiar with. On a board
consisting of 9 squares, arranged in a 3x3 matrix, players play alternately. On their turn, each player
writes a symbol in an empty square of their choice. One player uses the symbol X, the other uses the
symbol O. A player wins if they get three of their symbols lined up in a row, column or diagonal. If all
nine squares are filled up without this happening for either player, then the game is a draw. Write out a

set of rules, which if followed exactly, would allow me to play tic-tac-toe perfectly against all possible
opponents. That is, whether I play first or second, the rules should make me play the best possible move
for every situation that arises on the board.

Q8.
Assume the chance of it raining each day in Bangalore is 60%. You are equally unhappy when you get
wet and when you carry an umbrella on a non-rainy day. You are also equally happy when you have an
umbrella to protect you on a rainy day and when you are not burdened by an umbrella on a non-rainy
day. Then which of the following strategies will leave you most happy in the long run? (i) Always carry an
umbrella, (ii) Never carry an umbrella, (iii) Carry an umbrella 60% of the time, (iv) Carry an umbrella
50% of the time. Justify your answer. (You can assume that the 60% chance of rain is all the
information you have -- you cannot look out the window to see if rain clouds are appearing, how long it
rains is irrelevant, there is no correlation between it raining one day and it raining the next day, etc.)


Q9.
Consider a population of stem cells (P) which must go through an intermediate state (Q) before
ultimately differentiating into some final cell type (R). We model the PQ and QR transitions in
analogy to a chemical-kinetic model of three reactants (where p, q, r are concentrations; , are rate
constants), or a model of water flow between three tanks of identical size and shape (where p,q,r are
water levels, and ,reflect the efficiency of flow through the taps):






Such a system is represented by the following ordinary differential equations:

(i)

dp
dq
dr
= p, (ii)
= p q, (iii)
= q .
dt
dt
dt

Assume that initially all cells are in the undifferentiated state P, so that

p(0) = p0 , q(0) = 0, r(0) = 0 .


a.
For any time t, what is the value of p(t) + q(t) + r(t) ? [Hint: You dont need calculus to answer
this.]

b.
Solve for p(t). [Hint: radioactive decay.]

c.
Verify that the following function gives the number of cells in state Q:

q(t) = p0

et e t )
(

3

d.

[Hint: Differentiate this function wrt t and check that it satisfies Eq. (ii).]

Hint:

Suppose that the two rate constants are equal: = . Then it turns out that q(t) has the form:
q(t) = utewt . Find the values of u and w.

df (x)
f ( ) f ( )
. Substitute f (x) = ext , evaluate at x = .
= lim

dx

Again assuming = , at what time t will the number of cells in the intermediate pool be

e.
f.

greatest? (You may leave your answer in terms of u and w).

q(t), r(t) . Be as precise a
For p0 = 1, = = 1, sketch p(t),
s possible, label axes and all special
points.

Q10.

for different codons vary across tissues in humans. What do you expect will
The concentrations
of tRNAs
happen when the codon in present in an mRNA being translated but its associated tRNA is absent?
Explain how differing tRNA concentrations could lead to alternatively folded forms of a protein
translated from the same mRNA in two different tissues.


Q 11.
As can be seen from the figure, two complementary strands of DNA come
together to form a right handed antiparallel double helix.

(i) What are the possible benefits -with respect to function of DNA as
genetic material- of adopting this arrangement?
(ii) Protein factors are able to read the information resident in this
arrangement with exquisite specificity to bring about physiologically distinct
outcomes.
(iii) What different aspects of DNA in this form can be exploited by
protein factors to achieve specific binding?




Q 12. Using your biceps to lift a weight
The figure shows a simplified sketch of the biceps-elbow system. The
muscle-tendon unit represents the biceps, and m is the external
mass being lifted. Assume that bone #1 is attached to ground.
Dimensions are as indicated in the figure, in terms of `0.
When you lift a weight, like in the weights room in the gym, the
tension in the biceps' muscle-tendon unit creates a torque about the
elbow joint. The amount of torque generated depends on the lever
arm of the muscle-tendon unit about the joint centre of rotation.
This lever arm is called the `moment arm' of the muscle. The muscle-
tendon unit could `bow-string', causing the moment arm to depend
on posture. Calculate the moment arm of the biceps about the

elbow joint using the musculoskeletal geometry depicted in the figure. Find an expression for the
moment arm r as a function only of and `0.
Q13.
Our gut grows when we regularly increase food intake and shrink we regularly manage to diet severely.
Imagine a simplified gut with epithelial cells and smooth muscle. Keep in mind that food is digested,
nutrients taken up and sensed inside the animal. Write out a signalling mechanism that can detect levels
of food intake and respond to it by increasing gut size and reducing it on starvation. Link the specific
signalling pathways you choose, their ligands, receptors, output with the cell biology of secretion,
detection of systemic read-outs of nutrient levels and how this detection in turn feed back from the
body to the gut to regulate its size. You dont have to know the answer: You just need to present a
well-thought out credible answer.

Q14.
In a protein, a polypeptide chain can fold-back on itself: This common turn is called a beta-turn: How
many consecutive amino acid residues do you think are likely to be involved in a beta turn? Depending
on the numbers of consecutive amino-acid residues in a polypeptide chain involved in a turn the nature
of the turn can vary. Apart from the beta-turn, can you mention another turn, the numbers of amino-
acid residues involved and the nature of the turn?

Q15.
Stem cells can expand the size of tissues they make by switching from asymmetric cell division (linear
increase in cell number) to symmetric (exponential) division. Write down a plausible molecular
mechanism for asymmetric division where one daughter of a stem cell is a stem cell and the other a
daughter, which divides once and differentiates its two progeny. Write another plausible molecular
mechanism by which a stem cell gives rise to a daughter stem cell and a transit cell which then divides
to give two more transit cells, a process that continues for 6 rounds of symmetric transit cell division.
The amplified transit cells then divide once more to differentiate. You answer need not be based on
what we have learnt from research on this question: We just need a plausible molecular model that is
inspired by what we know of the cell biology of signalling, sub-cellular localization and transcriptional
response.

PART-1 Comprehensive Exam- June 2012

Part 1 (SECTION B) 2PM 6PM
Instruction: Attempt any ONE of the following questions

Projects should be framed using the following format:

Abstract 300 words or less
Introductory paragraph summarizing the background and framing the question
Hypotheses
Predictions
Experimental design framed to test the hypotheses
Analysis

Q1.
Reading material: Michael Hasselmos review (Trends in Cognitive Sciences Vol 3, 1999) on associative
memories and modulation by acetylcholine.

Project to design: Design a research project to test the acetylcholine gating theory for associative
memory in the hippocampal slice preparation. Your experiment can use any of the modern techniques:
2-photon calcium imaging, channel rhodopsin and optical stimulation, patch recording, transgenics and
so on.

Q2.
Reference: Senavirathne et al., Single-stranded DNA Scanning and Deamination by APOBEC3G at Single
Molecule Resolution. JBC, 2012
The enzyme APOBEC3G (Apo3G) catalyzes C-to-U deamination reactions in single-stranded DNA
(ssDNA), without requiring ATP or any small-molecule co-factors. It acts at the motif 5aaaCCCaaa3,
deaminating the final C. When there are multiple motifs on a single ssDNA molecule, Apo3G is known to
act processively: binding to a substrate strand, catalyzing multiple deamination reactions, then
unbinding. When short ssDNA strands are incubated with Apo3G, the final pattern of deamination
shows an asymmetry or polarity, favouring 5 motifs over 3 motifs. Our aim is to explain this polarity.

A. First, consider the hypothesis that Apo3G binds DNA, then scans unidirectionally, deaminating at all
motifs it passes until it unbinds. In which direction would the scanning have to occur? Is this model
tenable given the facts above?

B. Senavirathne et al. claim that their single-molecule experiments prove that Apo3G scans ssDNA bi-
directionally in a random fashion, based on the symmetric nature of their transition density plots (TDPs).
They state: All initial binding events stem from zero FRET on the y-axis, with most occurring at ~0.2
FRET, indicating that Apo3G binds preferentially away from the tethered 5-end. Strikingly the x-axis
and y-axis are completely symmetric in all TDPs, suggesting that Apo3G preferentially un-binds away
from the 5-end as well. Explain why this might be so.

C. Assuming that the scanning by Apo3G is indeed bidirectional with no bias, we must still explain the
polarized deamination pattern. The authors present a model involving the two DNA-binding domains (N-
terminal CD1 and C-terminal CD2) of Apo3G, only one of which is catalytically active (CD2). In their
model, Apo3G can bind DNA in two distinct orientations.



(i) Is this model sufficient to explain polarity?

(ii) What does the model predict for a long stretch of linear ssDNA?

(iii) What does the model predict for circular ssDNA?

(iv) What does the model predict for short linear double-stranded DNA?

D. Suggest further experiments to test the two-orientation hypothesis. Be specific, stating how each
proposed experiment would test aspects of the hypothesis.


Q3
How do humans throw at high speeds, yet achieve sufficient accuracy? Review the three papers
provided (Calvin, 1983, Chowdhary and Challis, 1999, Hore and Watts, 2011), and comment upon the
factors that govern accuracy of projectile release. What are the merits and limitations of the type of
analysis carried out by Calvin (1983) and by Chowdhary and Challis (1999)? Do the experimental findings
of Hore and Watts (2011) contradict the theoretical calculations before? Design an experimental
protocol, and propose a concurrent theoretical model, using control theory, to test the hypothesis
proposed by Hore and Watts (2011). How will you design an experiment to try and falsify your
theoretical model?

W H Calvin. A stones throw and its launch window: timing precision and its implications for language
and hominid brains. Journal of Theoretical Biology, 104(1):121135, 1983.
A G Chowdhary and J H Challis. Timing accuracy in human throwing. Journal of Theoretical Biology,
201(4):219229, 1999.
J Hore and S Watts. Skilled throwers use physics to time ball release to the nearest millisecond. Journal
of Neurophysiology, 106(4):20242033, 2011.

Q4.
Despite intense investigation, auditory transduction, the process by which sound is detected and
transduced by the sensory cells in the vertebrate inner ear remains incompletely understood. It is widely
accepted that auditory transduction is a process of mechanotransduction, which culminates with the
activation of ion channels on membranes of neurons in the inner ear. However despite several years of
analysis the identity of these ion channels remains unresolved.
Members of the TRP family of ion channels are implicated in transducing a range of stimuli in a
large number of species. The Corey lab has suggested that TRPA1 might be the mechanosensitive
channel involved in auditory transduction. Please read the primary research articles from the Corey lab
and discuss critically the evidence for/against the idea that TRPA1 channels are the final mediators of
auditory transduction. Suggest additional experiments that might be required to resolve this issue.


Q5.
1)Recent studies reveal genomic signatures of selection on genes involved in unique adaptations to
hypoxia (low oxygen levels) in humans that live in highlands, like the Tibetan Plateau (1, 2). Pikas are
lagomorphs (rabbit family) that live at high elevations (cold, hypoxic environments) or at high lattitudes
(like in Canada, cold enviornments). The tree below shows evolutionary relationships and distributions
of five pika species. Describe experiments to test whether (1) Pikas originally evolved adaptations to
hypoxia (2) they only have adaptations to cold environments and (3) they are adapted to cold and
hypoxia.

References:
1)
Sequencing of 50 Human Exomes Reveals Adaptation to High Altitude Yi, et al. Science 2 July
2010: Vol. 329 no. 5987 pp. 75-78. DOI:10.1126/science.1190371
2)
Genetic Evidence for High-Altitude Adaptation in Tibet Simonson et al., Science 2 July 2010: Vol.
329 no. 5987 pp. 72-75 DOI: 10.1126/science.1189406

Q6:
Pacific Salmon are born in fresh-water, migrate to the sea and return home to spawn. Read the
attached paper (Dittmann and Quinn, 1996; http://jeb.biologists.org/content/199/1/83) for a summary
of the process and what was understood some time. Today, as an experimental physicist interested in
biology, or as a behavioural biologist, what do you think the one key question to understand homing is?