Laminin-111 (Ln-1), an extracellular matrix (ECM) glycoprotein found in the basement membrane of mammary gland epithelia, is essential for lactation. In mammary epithelial cells (MECs), dystroglycan (Dg) is believed to be necessary for polymerization of laminin-111 into networks., thus we asked whether correct polymerization could compensate for Dg loss. Artificially polymerized laminin-111 and the laminin-glycoprotein mix Matrigel, both formed branching, spread networks with fractal dimensions from 1.7 to 1.8, whereas laminin-111 in neutral buffers formed small aggregates without fractal properties (a fractal dimension of 2). In Dg knockout cells, either polymerized laminin-111 or Matrigel readily attached to the cell surface, whereas aggregated laminin-111 did not. In contrast, polymerized and aggregated laminin-111 bound similarly to Dg knock-ins. Both polymerized laminin-111 and Matrigel promoted cell rounding, clustering, formation of tight junctions, and expression of milk proteins, whereas aggregated Ln-1 did not attach to cells or promote functional differentiation. These findings support that the microstructure of Ln-1 networks in the basement membrane regulates mammary epithelial cell function.

Historically, decline (or tristeza) has been a devastating disease of citrus caused by Citrus tristeza virus (CTV). Decline is a man-made disease based largely on propagation of sweet orange, grapefruit, and mandarins on the sour orange rootstock. In Florida, the major problem caused by CTV has been decline, since severe stem-pitting isolates are absent. Although this disease can be controlled by using alternative rootstocks, there are soils in which all other rootstock choices are less desirable in terms of fruit quality and yield. A major goal has been to develop measures that allow growers to use the sour orange rootstock in the presence of CTV. Florida has 2 predominant strains of CTV, a decline (T36) strain and a non-decline strain (T30). A first step was to map the viral determinant that induces decline. This was done by creating hybrids with T30 sequences substituted into T36 to identify sequences correlated with loss of decline symptoms. This project was delayed considerably because greenhouse assays to definitively assay decline symptoms did not work. In order to examine decline in field trees, a permit from the US Department of Agriculture Biotechnology Regulatory Service was obtained to test the recombinant-DNA-produced virus hybrids. This permit required that we test small trees inoculated with CTV at the time of planting. Under these conditions, those constructs that retained the T36 p23 and 3ʹ non-translated sequences induced greater amounts of stunting.

Laminin-111 (Ln-1), an extracellular matrix (ECM) glycoprotein found in the basement membrane of mammary gland epithelia, is essential for lactation. In mammary epithelial cells (MECs), dystroglycan (Dg) is believed to be necessary for polymerization of laminin-111 into networks., thus we asked whether correct polymerization could compensate for Dg loss. Artificially polymerized laminin-111 and the laminin-glycoprotein mix Matrigel, both formed branching, spread networks with fractal dimensions from 1.7 to 1.8, whereas laminin-111 in neutral buffers formed small aggregates without fractal properties (a fractal dimension of 2). In Dg knockout cells, either polymerized laminin-111 or Matrigel readily attached to the cell surface, whereas aggregated laminin-111 did not. In contrast, polymerized and aggregated laminin-111 bound similarly to Dg knock-ins. Both polymerized laminin-111 and Matrigel promoted cell rounding, clustering, formation of tight junctions, and expression of milk proteins, whereas aggregated Ln-1 did not attach to cells or promote functional differentiation. These findings support that the microstructure of Ln-1 networks in the basement membrane regulates mammary epithelial cell function.