Imaging Lipid Droplet Fusion and Growth - Methods in Cell Biology - page 247

Animals with Plin1 deficiency have a reduced adiposity and the accumulation of
smaller LDs in adipocytes due to the elevated basal lipolysis ( Castro-Chavez et al.,
2003; Martinez-Botas et al., 2000; Saha, Kojima, Martinez-Botas, Sunehag, & Chan,
2004; Tansey et al., 2001 ). In addition, Plin1 appears to induce LD clustering which
could be reversed by the phosphorylation of its serine 492 by PKA ( Marcinkiewicz,
Gauthier, Garcia, & Brasaemle, 2006 ).
Various mechanisms of LD growth
LD growth can be mediated by several distinct mechanisms. First, it has been shown
that several major enzymes for TAG synthesis are localized to LDs in various non-
adipocytes and locally produced TAG could be incorporated into the LD core and
promote LD enlargement ( Wilfling et al., 2013 ). For instance, overexpression of
LD-associated DGAT2 but not ER-associated DGAT1 promotes large LD formation
( Stone et al., 2009; Wilfling et al., 2013 ). Second, cytosolic LDs could stay in close
contact with ER where FIT1/2 and seipin/BSCL2 may regulate the TAG transfer
from ER to LD ( Fei et al., 2008; Gross, Zhan, & Silver, 2011; Kadereit et al.,
2008; Szymanski et al., 2007 ). The FATP-DGAT complex was also shown to pro-
mote LD expansion at the junction between LD and ER junction ( Xu et al., 2012 ). In
addition, ADRP and Rab18 might regulate TAG transfer from the ER to LD through
controlling the LD-ER contact ( Ozeki et al., 2005 ). LD growth on ER may require
specific ER domain/structure as depletion of atlastin or abolishment of atlastin
activity reduced LD sizes ( Klemm et al., 2013 ). Third, as LD surface is coated by
a monolayer of phospholipids, a phase coalescence, which resembles the SNARE-
mediated hemifusion, could directly result in the fusion between two LDs. The
phase coalescence is prevented by the joint efforts of both LD surface proteins and
phospholipids. Lack of phosphatidylcoline, a major component of LD surface phos-
pholipids ( Bartz et al., 2007 ), leads to excessive LD expansion to reduce the total
LD surface area ( Krahmer et al., 2011 ). The phase coalescence could also be induced
by exogenous chemical fusogens which may destabilize the LD surface structure
( Murphy, Martin, & Parton, 2010 ).
White adipocytes contain giant LDs that could occupy over 90% of intracellular
space. Although the three mechanisms mentioned above may all contribute to LD
growth in adipocytes, our recent work have demonstrated that fat-specific protein
27 (Fsp27/Cidec), a member of CIDE family proteins, plays a major role in the gen-
eration of unilocular LD in adipocytes by mediating atypical LD fusion through a
directional neutral lipid transfer at LD-LD contact site (LDCS) ( Gong et al.,
2011 ). Fsp27 is highly expressed in adipose tissue and its deficiency leads to the
accumulation of smaller and multilocular LDs in white adipocytes and drastically
reduced lipid storage ( Gong, Sun, & Li, 2009; Nishino et al., 2008; Toh et al.,
2008 ). In contrast, ectopic expression of Fsp27 results in an increase in LD sizes
( Jambunathan, Yin, Khan, Tamori, & Puri, 2011; Keller et al., 2008; Liu et al.,
2009; Puri et al., 2007, 2008 ). Mechanistically, when two or more LDs are in close
apposition, Fsp27 is rapidly accumulated and enriched at the LDCS where it may
create a putative pore allowing neutral lipid exchange between connected LDs.
The final step of Fsp27-mediated LD fusion is dependent on the internal pressure