The Ldb1 Transcriptional Co‐Regulator is Required for Establishment, Development, and Survival of the Pancreatic Endocrine Lineage

Abstract

Pancreatic islets are comprised of hormone secreting cell types that are vital regulators of glucose metabolism. Specifically, the pancreatic β cell is indispensable for glucose control and its dysfunction is central to diabetes mellitus. β cell development is regulated by transcription factor (TF) cascades, mediating differentiation of progenitors into mature insulin producing β cells. Our prior studies show that TF, Islet1 (Isl1), interacts with a transcriptional co‐regulator, Ldb1, to regulate β cell maturation from embryonic day (E) 18.5 onward. However, Ldb1 is also expressed in early stages of pancreas development, before Isl1 is present (as early as E10.5). The earlier Ldb1+ cell types include Pdx1+ multipotent progenitor cells (MPCs) and endocrine progenitors expressing the TF Neurogenin3 (Ngn3). MPC progeny will populate the entire pancreas (endocrine and exocrine), while Ngn3+ endocrine progenitors (Isl1−) are fated to become islet cells. Our hypothesis is that Ldb1 has Isl1 independent roles in maintaining progenitor identity in these requisite populations during early pancreatic development. To test this we generated a whole pancreas knockout of Ldb1 (Ldb1Δpanc) and observed severe developmental and postnatal phenotypes. At E13.5, Ldb1Δpanc mice exhibit disorganized progenitor pools, suggesting early defects in endocrine identity. At E15.5, Ldb1Δpanc mice had a significant reduction of Ngn3+ progenitors and Pdx1HI immunoreactivity, a mark of presumptive β cells. Ldb1Δpanc mice die by postnatal day 7 (P7) with severe hyperglycemia and hypoinsulinemia due to drastic islet hormone cell reduction. Interestingly, total pancreatic mass remained unchanged in Ldb1Δpanc neonates, suggesting that Ldb1 impacts are islet specific in the pancreas. Considering these observations, we generated a new model of Ldb1 loss specifically in Ngn3+ islet progenitors, termed Ldb1Δendo. We confirmed loss of Ldb1 in endocrine clusters and observed postnatal hyperglycemia, with a reduction of islet cells in neonates, similar to that seen in Ldb1Δpanc mice. Chromatin immunoprecipitation in vivo and in vitro highlights that Ldb1 imparts control on Pdx1 through occupation of the Pdx1 Area I–II regulatory domains, and that Ngn3 control also occurs via direct occupation. We are now further assessing the developmental phenotype, examining markers of proliferation, apoptosis, and altered cell identity through lineage tracing in the Ldb1Δendo model. Concomitantly, our published work revealed that Ldb1 and Isl1 interact in complex with the single stranded DNA binding protein co‐regulator, SSBP3, which helps stabilize this transcriptional complex. To examine the functional relationship between SSBP3, Isl1, and Ldb1 in vivo, we developed a new SSBP3 floxed mouse model in order to generate pancreas wide loss of SSBP3 (SSBP3Δpanc). We observed preliminary reductions in key islet mRNAs along with hyperglycemia in neonates at P1. Our work provides insight into the transcriptional complexes dictating how islet progenitors adopt their cell fate. Enhancing our understanding of this differentiation process can contribute to therapeutic β and islet cell generation strategies to help the growing diabetic population.Support or Funding InformationNIH‐NIDDK R01DK111483‐01NIH‐F31DK120217NIH‐NIGMS T32GM008111