Downregulation of Protein tyrosine phosphatase PTP-BL represses adipogenesis

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Time-dependent modulation of intracellular signaling molecules and sequential induction of transcriptional regulators that generate marked changes in gene expression are essential for the differentiation of pre-adipocytes into adipocytes (Rangwala and Lazar, 2000) (Koutnikova and Auwerx, 2001). The transcriptional regulation of adipogenesis is relatively well characterized, with PPARg and C/EBPa as two master regulators that control adipogenic genes (Nedergaard et al., 2005). However, the factors that influence pre-adipocyte determination remain poorly understood. Here we demonstrate that the mouse protein tyrosine phosphatase PTP-BL is strongly expressed in white adipose tissue and its expression levels are upregulated during proliferation and differentiation of 3T3-L1 pre-adipocytes, suggesting a role for PTP-BL in adipose tissue biology. Indeed, we found that PTP-BL plays a key role in 3T3-L1 adipocyte differentiation. We show that blocking the up-regulation of PTP-BL expression in the early phases of adipogenesis was associated with a dramatic decrease in adipogenic gene expression and lipid accumulation in 3T3-L1 adipocytes.

A number of diverse functions have been described to PTP-BL, given the varied proteins that it has been shown to interact with. For example, PTP-BL has been suggested to act as a scaffolding protein in the regulation of the cytoskeleton (Erdmann, 2003), to dephosphorylate EphrinB and thus regulate various developmental processes (Palmer et al., 2002), and to be involved in the regulation of cytokinesis (Herrmann et al., 2003). Reports based on the characterization of mice that lack PTP-BL phosphatase activity revealed a phenotype of impairment in motor nerve repair (Wansink et al., 2004) and the involvement of PTP-BL in both retinal ganglion cell neurite initiation and survival of activated retinal glia (Lorber et al., 2005). Recently, a published study on mutant mice that are completely devoid of PTP-BL did not report phenotypic consequences or an effect on adipogenesis and solely focalized on haematopoietic cell lineages (Nakahira et al., 2007) which were previously shown to express the phosphatase (Gjorloff-Wingren et al., 2000). For the human homologue of PTP-BL, PTPL1, a role in apoptosis has been proposed, but this is still a matter of debate and depending on Fas pathway effectiveness and PTPL1 expression level (Sato et al., 1995) (Abaan et al., 2005; Bompard et al., 2002; Cuppen et al., 2000; Miyazaki et al., 2006; Toretsky et al., 1997; Wieckowski et al., 2007). Our own studies have shown that PTPL1 is necessary (Bompard et al., 2002) and sufficient (Dromard et al., 2007) for the early inhibition of the IRS-1/PI3K/Akt pathway through IRS-1 dephosphorylation.

The insulin/IGF-1/IRS signaling pathway has an important role in the regulation of genes that are involved in multiple early adipogenic events in brown pre-adipocytes (Tseng et al., 2005). Activation of Akt is considered to be important for adipogenesis (Fasshauer et al., 2001; Sakaue et al., 1998; Xia and Serrero, 1999; Xu and Liao, 2004; Yoshiga et al., 2007). The Akt signal cascade appears to induce or activate PPARg and C/EBP during the induction of 3T3-L1 adipocyte differentiation (Kim and Chen, 2004; Kortum et al., 2005; Lazar, 2005; Nedergaard et al., 2005). Recently, PPARg2 expression was shown to be negatively regulated by Foxo-1, through binding to the PPARg promoter and inhibition of transcription. (Armoni et al., 2006). Insulin induces phosphorylation and nuclear export of Foxo-1 through Akt. Therefore, the IR/IGF-1R-IRS-PI3-kinase-Akt pathway promotes PPARg expression. Surprisingly, we found that PTP-BL knock-down had no apparent effect on IRS-1 tyrosine phosphorylation and Akt activation in 3T3-L1 pre-adipocytes. This may be indicative for compensatory actions from other PTPs present in these cells but may also point to a regulatory mechanism controlling PTP-BL activity as encountered during oocyte meiotic maturation (Nedachi and Conti, 2004). Still, PTP-BL was found to be required during the early phase of adipocyte differentiation that directly follows the clonal expansion phase. Interestingly, we could demonstrate that PTP-BL phosphatase activity is not required for this effect on adipocyte differentiation, in line with the absence of an effect of PTP-BL on the IRS-1/Akt pathway. Indeed, MEFs (C57BL/6) expressing a PTP-BL protein lacking its PTP domain differentiated into the adipogenic lineage as efficiently as wild type cells. This is in accordance with the fact that PTP-BL^DP/DP mice, that lack the PTP-BL catalytic domain, do not display any abnormality in adipose tissue development when bred onto a C57BL/6 genetic background (Wansink et al., 2004). In that light, it will be interesting to learn whether any adipose tissue abnormalities become apparent in the full PTP-BL knock-out mice that were recently generated (Nakahira et al., 2007).

PTP-BL and PTPL1 represent the largest mammalian intracellular PTP in mice and human, respectively, and possess at least seven potential protein-protein interaction domains; a kinase noncatalytic C-lobe (KIND) domain, a Four-point-one-Ezrin-Radixin-Moesin (FERM) domain, and five PSD-95-Drosophila discs large-Zonula occludens (PDZ) domains. Many potential interacting proteins have been identified (Erdmann, 2003). The contribution of PTP-BL to the adipocyte differentiation process seems to be reflected by its potency to orchestrate the composition and dynamics of large protein machines through these many protein interaction modules rather than by its enzymatic activity. The precise molecular mechanism by which PTP-BL participates in adipogenesis, however, remains to be uncovered.

Elucidation of PTP-BL’s working mechanism as an adipogenic factor might result in new therapeutic strategies for the treatment of obesity that will involve the abrogation of PTP-BL protein complexes rather than inhibition of its enzymatic activity.