The role of nuclear lipid signalling in adipocyte differentiation

Abstract

The differentiation of preadipocytes into adipocytes is one of the most intensely studied biological processes and has led to the understanding of a number of signaling pathways involved in cell growth and differentiation. The nuclear phosphatidyl inositol (pl) signalling pathway, centred around the enzyme phospholipase C (PLC) βI has been demonstrated to be involved in a number of cellular processes such as mitogenesis and differentiation. Recently components of another lipid signalling pathway, that involving prostaglandins, have also been shown to localise at or within the nucleus. To date, there have been no reports demonstrating the role of either of these nuclear pathways during the differentiation of adipocytes. The 3T3-L1 preadipocyte cell line was used to study the role of nuclear pI and prostaglandin lipid signalling in adipocyte differentiation. In Chapter 3 data is presented to show that two phases of nuclear PLC βl activity occur during adipocyte differentiation. The first phase occurs within minutes of differentiation being induced, is regulated by the actions of extracellular signal-related kinase (ERK) l/2 and protein kinase c (pKC) α, and is involved in the mitotic clonal expansion phase of differentiation. The second phase of activity occurs from day 2 of differentiation, is regulated independently of ERK 1/2 and pKC α and is involved in the regulation of the cell cycle factors cyclin D3 and cyclin-dependent kinase (cdk) 4. In Chapter 4 I investigate the nuclear localisation of prostaglandin synthases, how their localisation is altered by differentiation, and the role of these enzymes in adipocyte differentiation is characterized through inhibitor studies. Cytosolic phospholipase A2 (cPLA2) cyclooxygenase (COX) 1 and 2, microsomal prostaglandin E2 synthase (mPGES), prostacyclin synthase (PGIS), lipocalin prostaglandin D2 synthase (L-PGDS) are localised to the nuclear membrane and their localisation changes during differentiation. The COX 1 and2 enzymes also localise within the nucleus and this COX 1 localisation appears to be cell cycle dependent. COX 2 is functionally coupled to mPGES and PGIS and plays a role in adipocyte differentiation as inhibition of COX 2 with a specific inhibitor (NS 398) completely blocks adipocyte differentiation. COX 1 is functionally coupled with L-PGDS and appears to be involved in lipid metabolism as the inhibition of COX 1 with a specific inhinitor (SC 560) partially blocks lipogenesis. In Chapter 5, I investigate the localisation of prostaglandin receptors in the differentiation process. 3T3- L1 cells express the prostaglandin E2 (EP4) receptor, prostaglandin F2α (FP) receptor and the prostaglandin D2 (DP) receptor but these do not have a nuclear localisation. Activation of the DP receptor is involved in lipid metabolism as the DP receptor antagonist, BW A868C, partially inhibits lipogenesis. lnternalisation of prostaglandins appears to be necessary for differentiation to occur fully as treatment of differentiating 3T3-L1 preadipocytes with the anionic transport inhibitor bromocresol green (BCG) partially inhibits carbaprostacyclin (cPGI2) induced differentiation and completely inhibits that caused by cyclopentenone prostaglandins. Thus, nuclear lipid signalling pathways appear to play a role in the process of adipocyte differentiation and a proper understanding of these pathways may lead to a better understanding of fat cell development and the role it plays in obesity and its related diseases.