Temporal and spatial expression of laminin (LN) isoforms is critical to glomerular development and contributes to altered cell function in diabetic nephropathy and other progressive renal diseases. Yet, little is known about the mechanisms that regulate LN isoform expression. We propose that insulin-like growth factor binding protein-5 (IGFBP-5) leads to formation of a filamin-based nuclear shuttle that binds transcription factors (e.g. sox9, GKLF, Sp1, Smad) and transcriptional co-activators (e.g., IGFBP-5 and LIM domain proteins such as FHL2) and regulates laminin (LN) isoform expression in mesangial cells (MC). This hypothesis suggests a novel mechanism whereby perturbations in the cell-matrix interface regulate gene expression. This hypothesis will be examined by:
Specific Aim 1 : To characterize the generation and composition of the filamin nuclear shuttle following treatment of MC with IGFBP-5. First, we will demonstrate that a fragment of filamin forms and translocates to the nucleus. Second, we will characterize the transcription factors that are recruited to the complex; and finally, we will evaluate the role of transcriptional co-activators, LIM domain protein FHL2 and IGFBP-5, in the complex.
Specific Aim 2 : To demonstrate that nuclear accumulation of filamin is required for IGFBP-5-mediated changes in LN gene expression. First, we will demonstrate that filamin is required for IGFBP-5-mediated effects on LN gene expression. Second, we will show that the filamin shuttle is recruited to LN gene loci where it specifically modifies LN gene transcription.
Specific Aim 3 : To define the mechanisms whereby the filamin nuclear shuttle activates LN gene transcription. First, we will determine if FLN interacts directly with the LN promoter and if it drives transcription. Second, we will define the elements in the shuttle that are required for transcriptional control. These studies will define the role of a filamin-based nuclear shuttle in mediating the effects of IGFBP-5 on laminin gene expression. Understanding transcriptional control of LN isoform expression will add new insights into the role that LN plays in normal glomerulogenesis and that becomes disordered in glomerular disease. ? ? ?
| Zhang, Jiong; Hansen, Kim M; Pippin, Jeffrey W et al. (2012) De novo expression of podocyte proteins in parietal epithelial cells in experimental aging nephropathy. Am J Physiol Renal Physiol 302:F571-80 |
| Schaeffer, Valerie; Hansen, Kim M; Morris, David R et al. (2012) RNA-binding protein IGF2BP2/IMP2 is required for laminin-ýý2 mRNA translation and is modulated by glucose concentration. Am J Physiol Renal Physiol 303:F75-82 |
