Biology Reference
In-Depth Information
A complete understanding of the dynamic change of lipid transfer rate as a function
of donor and acceptor LD sizes still await large volume of high precision live
cell imaging of LD fusion events and mathematical modeling. Currently, the lipid
transfer rate is quantified as the average lipid transfer rate of the entire LD fusion
process. Thus, similarly sized LD pairs are used for quantitative comparison.
The initial size of donor (smaller) LD (in diameter) is measured as the average of
three consecutive frames when the ring circle of Fsp27-GFP signal on LD surface is
clearly visible. The end point of the completion of lipid transfer is defined as the
donor LD showing a punctate structure attaching to the surface of large LD in five
consecutive frames. We calculated the lipid transfer rate by dividing the volume of
the donor LD with the duration of the lipid transfer. Using this method, we have
shown that Plin1 significantly accelerates the rate of Fsp27-mediated lipid transfer
and LD fusion in 3T3-L1 preadipocytes (
Sun et al., 2013
). As shown in
Fig. 14.3
, the
LD fusion pair in cell expressing Fsp27-GFP along (upper panel) contained a donor
LD of 2.6
m
3
min
diameter. The LD fusion event took 80 min with an average lipid transfer rate of
m
m in diameter (9.2
m
in volume) and an acceptor LD of 5.2
m
Fsp27-GFP
0 min
10.5 min
20 min
34 min
46.5 min
54.5 min
65 min
80 min
0.3
***
0.2
0.1
Fsp27-GFP + HA-Plin1
0.0
0 min
11 min
21.5 min
37 min
47 min
49 min
56 min
65 min
FIGURE 14.3
Representative time-lapse images showing that Fsp27 promotes lipid transfer and Plin1
enhances Fsp27when coexpressed in 3T3-L1 preadipocytes. Red asterisks and yellow
arrowheads represent the accepter and donor LDs, respectively. Bars, 4
m (left panels). The
net neutral lipid transfer rate shows that Plin1 significantly accelerates Fsp27-mediated
lipid transfer and LD fusion (***p<0.001).
m
