BIOL-100B late. Associated Reading: Golgi maturation visualized in living

BIOL-100B Advanced Cell Biology Research Article Assignment
1 Due: On LATTE, 11:59pm January 21st, 2018. Not accepted late. Associated
Reading: Golgi maturation visualized in living yeast. Losev E, Reinke CA,
Jellen J, Strongin DE, Bevis BJ, Glick BS. Nature. 2006 Jun
22;441(7096):1002-6. Epub 2006 May 14. Paper and supplements are available more
conveniently on LATTE. Note: Questions 1-6 are worth 1 point and will be graded
individually on “good faith effort.” This means that you do not need to have
the correct answer to get full credit, but you do need to demonstrate that you
carefully read the paper and tried to answer the question correctly. Any of
these questions could appear on any of the tests. If they do, they will be graded
for correctness. Question 7 is optional and will not be graded. How to submit:
This assignment must be submitted online via Latte, as a pdf or text document.

1. What is the big
question of this paper? Not, what is this paper about, but what problem is this
entire field trying to solve?

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What is the nature of the early (cis, medial) and late
(trans, TGN) cisternae and how do they mature?

2. Summarize the
background in five sentences or less. Bullet points are ok.

·       
Determine florescence of early Golgi proteins
tagged with GFP and late Golgi protein with DsRed using S. cerevisiae

·       
The Golgi Dynamics were examined using 4D
microscopy, using markers Sec7 for late cisternae and VRG4 for early cisternae.

·       
The Maturation Mechanism was consistent through
carboxypeptidase and visualized cisternal maturation by tagging both markers at
the same time – GFP withVrg4 and Sec7 with DsRed.

·       
 Future
experiments will test the association between maturing cisternae and secretory
cargo proteins

3. What is(are) the
specific question(s) of this article (i.e. what exactly are the authors trying
to answer with their research)?

·       
Does the rate of cisternal maturation match the
rate of protein transport?              

o  
Was the composition of Golgi-resident proteins
changed over time when visualized individually?

o  
How are they going to use light microscopy so
that they can distinguish individual cisternae?

o  
How were they going to fluorescently tag Golgi
marker proteins that were highly expressed while not affecting the organelle?

4. What experimental
system and approach are they using?

3D time-lapse fluorescence microscopy of yeast S. cerevisiae

5. Explain Figure 4
and S6:

a. Describe the experiment.

Radioactive pulse-chase
measurement with rapidly secreted protein in yeast, Alpha-factor was used to
determine whether the experiments done were accurate and the maturation matched
the rate of secretory traffic. In the Alpha- factor, the core slowly disappeared
at time went on, and in the Carboxypeptidase Y, the gel shows that as time went
on, the mature bands showed darker and darker bands.

 

b. What question(s) do(es) this experiment
help the experimenter answer?

 

Is there a speed
difference between Golgi-to-cell surface transport and intra-Golgi transport?

 

c. What results are shown in this figure?
Be sure to explain the axes, what is being depicted in the associated panels in
the supplemental figure, and the meaning of any variables.

 

A.      The
Band intensity (% of max) is the y axes and it is describing the band on figure
S6 and how the core at .5 min was at 100% and then decreased to 0% by minute
10. And the maturity in figure S6 increased to 100% intensity by minute 14
(x-axes is chase time in minutes).

B.       For
Carboxypeptidase Y, even though is it less accurate, showed a similar graph
where band intensity of p1 is 100% from 0-2min, and maturity reached around 80%
by minute 40.

For both graphs
the red line indicated ER-localized core and p1, and the blue dotted line
indicated halftime disappearance and vacuole-localized mature CPY.

d. How do the results contribute to
answering the question(s) you outlined in “B”?

These results indicate
that Golgi-to-cell-surface transport is in fact several times faster than intra
Golgi transport which goes back to the original question of the paper which was
the rate of cisternal maturation.

 

6. Do the results of
this paper help the answer the big question you outlined in “1?” Why or why
not?

Yes, but to a degree, it answers part of the big question of
the rate of cisternal maturation, which is a portion of the bigger question,
but more research needs to be done to actually answer the full question. But
this study will help forward the future research.

7. Is there anything
you’d like the presenter(s) to know or address during their presentation? For
example, questions about terminology, methods, or results. Great videos,
diagrams, or other resources you may have found. (optional)

I thought P1 and p2 were poorly explained, and
think it would be interesting to maybe address more what they are and then
reanalyze