Obesity continues to escalate as a significant public health problem and as the leading preventable cause of death. Genetic, environmental, behavioral, and socioeconomic factors cause excess weight gain and obesity. Increased proliferation and differentiation of pre-adipocytes to mature adipocytes (adipogenesis) within the fat tissues are central to obesity. Previous studies focused extensively on the transcriptional controls which maintain the adipocyte phenotype. However, the underlying molecular mechanisms that promote determination of primitive mesenchymal precursor cells to the adipose phenotype are still unknown. Our long-term goal is to elucidate the cellular and molecular mechanisms underlying adipogenesis. In pursuit of this goal, our objective is to determine how alternative splicing of apoptotic genes govern the differentiation and maturation of pre-adipocytes. It is well documented that pre-adipocytes undergo apoptosis while mature adipocytes fail to do so. Based on preliminary data, our central hypothesis is that there is a distinct shift in the expression of genes involved in apoptosis from a pro-apoptotic to a pro-survival pathway via alternative splicing controlling terminal adipocyte differentiation which ultimately increases the adipose number and mass leading to obesity. The significance of the proposed research is that once we establish the molecular mechanisms of adipogenesis which result in increased survival of adipocytes, we can manipulate targets of the apoptosis pathway to devise new and innovative approaches to prevent or reverse weight gain and obesity. We propose three specific aims.
In Aim 1, we will elucidate whether terminal adipocyte differentiation is accompanied by alternative expression of apoptosis genes. Pre-adipocytes undergo apoptosis unlike mature adipocytes. Our data demonstrate that during adipocyte differentiation between days 4-6, expression of Bcl2 and Bcl-x, caspase 9 and PKC4 alternatively spliced variants change to its pro-survival variants. Using overexpression and knockdown experiments, we will elucidate the link between these genes, adipogenesis and apoptosis.
Aim 2 is designed to determine the role of the signaling kinase PKC4II as an anti-adipogenic mediator. PKC4II, a serine/threonine protein kinase promotes cell survival. Natural compounds such as resveratrol and curcumin exhibit anti-adipogenic activities and promote apoptosis in adipocytes. Our data shows that they inhibit PKC4II expression. Our data further points to PKC4II as the signaling kinase upstream of Bcl2-mediated survival pathway. We propose to identify the intracellular target which is central to pro-survival pathways in adipocytes that may be modulated to induce anti-adipogenic effects.
In Aim 3 the physiological relevance of this hypothesis will be assessed determining the expression levels of these apoptosis genes in diet-induced obese mouse model (C57BL/6) fed a high-fat diet complemented with the natural compounds or a PKC4II-specific inhibitor. By showing that alternative splicing of apoptotic genes modulate differentiation and maturation of pre-adipocytes, the proposed studies will establish a new paradigm for adipogenesis and reveal new targets for treatment of obesity.
The obesity epidemic is increasing at an alarming rate in USA. In the largest obesity prevalence study to date in VA, it was found that amongst veterans receiving healthcare at VA facilities overweight and obesity were at a significantly higher prevalence than the general population. In 2005, Dr. Perlin (then-acting Under Secretary of Health) announced the MOVE! (Managing weight and/or obesity for veterans everywhere) program to combat obesity and its related diseases. This program is given the highest priority as it affects current treatments and future demand for VA healthcare services. The proposed studies are expected to have important therapeutic implications as we expect to identify new targets for interventions to prevent or reverse weight gain and thereby impede the rise of obesity-related morbidities.