The mechanism of LPL regulation is complex, and we have focused on the translational regulation of LPL, which occurs in response to diabetes, and in response to hormones such as thyroid hormone and epinephrine. LPL translation is controlled by an RNA binding complex that forms in response to epinephrine and binds to the 3'UTR of the LPL mRNA, resulting in an inhibition of LPL translation. 1 component of this complex is a member of the A-Kinase Anchoring Protein (AKAP) family, which tethers PKA to specific cellular sites. AKAP121 contains a consensus KH RNA binding region and is a component of the RNA binding complex. Additional studies have characterized the region on the LPL mRNA that binds the AKAP-PKA complex, and demonstrated that transgenic mice expressing LPL without the proximal 3'UTR upregulate LPL at the translational level. Therefore, we plan to examine LPL translation regulation by studying both the RNA binding complex, and the LPL mRNA. Hypothesis 1. LPL translation is regulated by an RNA binding complex which interacts with a specific motif on the LPL 3'UTR. This RNA binding complex depends on specific sequence elements of PKA, AKAP149, and possibly other components.
Aim 1. Examine AKAP149/121 interactions with other elements of the PKA complex.
Aim 2. Determine whether AKAP targeting to ER or mitochondria is important in LPL translation inhibition.
Aim 3. Will the introduction of AKAP149 confer LPL translational inhibition by PKA in a non-adipose cell? Aim 4. Is the increase in LPL that accompanies hypothyroidism due to changes in AKAP expression? Aim 5. Identify LPL 3'UTR sequence elements that alter the interaction with the RNA binding complex. Hypothesis 2. By preventing the interaction between the RNA binding complex with the LPL 3'UTR, LPL will become unresponsive to epinephrine, and adipose tissue will accumulate more lipid, diverting lipid from other insulin target organs, yielding improved insulin sensitivity.
Aim 6. Examine the phenotype of a transgenic mouse that expresses LPL lacking the proximal 3'UTR. Will this cause lipid partitioning into adipose tissue, and improve insulin sensitivity? Aim 7. Examine insulin sensitivity and other phenotypic features of d-AKAP1 knockout mice. Relevance. LPL is a central enzyme in lipid metabolism: the enzyme is expressed by adipocytes and muscle cells, and hydrolyzes the triglyceride core of plasma VLDL and chylomicrons. LPL is an important adipogenic enzyme in adipose tissue, and abnormalities in LPL result in inefficient lipoprotein metabolism, leading to atherosclerosis, especially in patients with diabetes. ? ? ?
