At least three major mitotic processes are regulated by the PP2A-B56 phosphatase complex: the Spindle Assembly Checkpoint (SAC), kinetochore-microtubule attachments and sister chromatid cohesion. We show here that these key functions of PP2A-B56, which require its localization to either the kinetochore or centromere, are split between distinct subsets of B56 isoforms. PP2A-B56γ and PP2A-B56δ localize to the outer kinetochore (via BUBR1), whereas PP2A-B56α and PP2A-B56ε localize to the centromere (via Sgo2). The differential localization observed is due to a difference in affinity for the receptors: PP2A-B56γ has a reduced affinity for Sgo2 compared to PP2A-B56α and, vice versa, PP2A-B56α has a reduced affinity for BUBR1 compared to PP2A-B56γ. Given that the known binding interfaces for both BUBR1 and Sgo2 are highly conserved in all B56 isoform, we generated a series of chimeras between B56α and B56γ to uncover isoform specific interactions. This led to the identifications of two distinct regions within B56α and B56γ that regulate the binding to Sgo2 and BUBR1. Furthermore, site directed mutagenesis has revealed that proper holoenzyme assembly has a role in regulating the localization of B56: it is needed for centromeric accumulation and it interferes with kinetochore accumulation of B56α. We will present a model to explain how this differential localization could be linked to post-translational modifications of PP2AC. Together, these results help to clarify how individual PP2A-B56 isoforms achieve subcellular specificity during mitosis.

If y₁, y₂, ... , y _p represent vectors of independent observations, the generalized multivariate regression model is of the form y_j = X_1j β_1j + X_2j β_2j + ε_j , j = 1, 2, …, p, where X_1j and X_2j are general linear model regression matrices, β_1j and β_2j are vectors of unknown coefficients, and the ε_j are error vectors such that cov(ε_i,ε_j) = σ_ijI. If X_1j = X₁ and X_2j = X₂ , j = 1, 2, …, p, the above is a standard multivariate regression model . Insofar as can be determined, the true relationship between the design variables and a response n_j is n_j = x_1j^’ β_1j + x_2j^’ β_2j where x_1j^’ x_2j^’ are typical row vectors in the matrices X_1j and X_2j. For x_j^* = [x_1j^’, x_2j^’] and β_j^* = [ß_1j^’, β_2j^’], the n_j are to be estimated either by ŷ_j = x_1j^’β̂_1j or ŷ_j* = x_j^*’ β̂_j^* where β̂_1j and β̂_j^* are the least squares estimators of β_1j and β_j^* obtained from the full multivariate regression model. The estimators for the n_j are determined by a test of the hypothesis H_o: J₁ ≤ J₂ where J₁ and J₂ denote the integrated mean squared errors of a linear combination of the ŷ_j and ŷ_j* respectively. Rejection of H_o results in selection of the ŷ_j*; otherwise the ŷ_j are chosen. A test statistic is developed to test H_o with consideration extending to several important special cases. Distinctions are drawn between the preliminary test estimator constructed around H_o, and that based on the usual hypothesis β_2j = 0, j = 1, 2, ..., p. Under the assumption of error normality, an approximation to the distribution of the test statistic is developed in order to determine type I and type II error probabilities. An explicit expression for J_o, the integrated mean squared error of the preliminary test estimator, is obtained, and difficulties in its evaluation are discussed. An estimator of J_o is presented along with a special case in which J_o can be evaluated exactly. Graphical comparisons are made on the relative performance of the estimators based on H_o , and those constructed around the standard hypothesis. An operating range of type I error probabilities is also discussed. / Ph. D.

The specific objectives of this study were: 1) to develop a price forecasting equation for feeder cattle and 2) to examine the possibility of reducing the variance in income and raising the level of revenue relative to that which would be achieved under the common practice of consistently selling feeder cattle in the cash market. To these ends, a structural model for the feeder cattle market in the United States was developed in order to isolate the important variables to be used in the construction of a quarterly price forecasting model. Time series methods described by Box and Jenkins were employed in the construction of the forecasting equations. Selected strategies were developed which incorporated trading in the futures market for feeder cattle based on the price forecasts. The trade-off between mean income and the mean square error of income is indicated by a test, within the range of the data base, of the various strategies for the volatile years 1972-1975. / Ph. D.

With the advent of the "Space Shuttle" concept, it has become necessary to study the effects of heat transfer on the aerodynamic and boundary layer characteristics of a heated delta wing. Thus, a symmetrical 60° delta wing was tested up to twice freestream temperature in the Virginia Tech 6' x 6' stability wind tunnel. Summaries of the characteristics of the flow over the unheated wing and the theoretical effects of heat transfer are included. It has been found that heat transfer effects on the wing's aerodynamic characteristics are negligible at angles of attack up to one-third of the maximum lift angle. Beyond this, lift and pitching moment show a very small decrease and increase, respectively, up to maximum lift while drag increases 15 percent to 25 percent at maximum lift. Further increases in drag occur when the wing is yawed. No decrease in stalling angle of attack with heating is found for all yaw angles. / Master of Science

A total of 62 solid waste disposal sites were documented and evaluated in the 18 county study area of southwestern Virginia. The solid waste disposal operations encompassed a combined area in excess of 458.7 ha. An estimated 32 workers and 40 pieces of heavy equipment were utilized for the disposal of the waste at the various dumps, landfills, and sanitary landfills. The largest number of disposal sites were operated as dumps (62%), while only 14% of the sites qualified as sanitary landfills. The dump type of operation was characterized by: erosion, odor, leachate, minimum compaction, burning, lack of final or daily cover, and limited soil, financing, cover material, personnel, equipment, and planning. Generally, the waste was deposited at a remote or obscure site and left unattended. The landfill form of waste disposal, which occurred in 17% of the sites, required the construction of a trench for the disposal of the waste material which was later covered. The landfill form of waste disposal was characterized by shallow soils, limited cover material, steep slopes, and/or underground springs. The sanitary landfills were located in areas of deep soils, with moderate slopes, and showed an absence of interacting water. These sites demonstrated proper management of the resources and limited alteration of the surrounding area. / Master of Science

Thesis (M.S.)--Massachusetts Institute of Technology, Dept. of Architecture; and, (M.S.)--Massachusetts Institute of Technology, Dept. of Mechanical Engineering, 1986. / Typescript (photocopy). / Includes bibliographical references (v. 2, leaves 288-292) / A study of the energy system at the Massachusetts State Transportation Building was conducted. This innovative energy system utilizes internal-source heat pumps and a water thermal storage system to provide building heating and cooling. The potential benefits of this type of system include both energy savings and operating and equipment cost savings when compared to more conventional building heating and cooling systems. The study involved monitoring of equipment performance, computer simulation of the building energy system dynamics, and analysis of actual and modelled system efficiency. It was found that the building is presently operating as a 'low energy' building, despite a number of factors which have limited the heat pump system's capability to entirely meet winter heating requirements. Significant additional operation efficiency and cost savings are potentially available if a variety of measures are undertaken, including: stratification of the thermal storage system, utilization of demand management controls, and increased lighting system efficiency. / by Abbe Ellen Bjorklund. / M.S.

Dictyostelium discoideum is a soil dwelling amoeba which has been widely used as a model organism to study cellular processes such as signal transduction, chemotaxis, endocytosis and exocytosis. The process of phagocytosis in Dicytostelium is largely comparable to that of neutrophils and macrophages in the mammalian system. Neutrophils and macrophages are cells of the innate immune system and they engulf infectious bacteria through phagocytosis. Dictyostelium cells uptake yeast and bacteria for their nutrition through phagocytosis, which is an actin dependent mechanism and is a target of multiple signaling inputs. Recent studies have uncovered different proteins involved in the signaling of particle and further studies are required to decipher the intricate mechanism leading to the F-actin rearrangement. Two of the proteins have previously known to be involved in the pathways regulating the F- actin rearrangement name PP2A phosphatase and LKB1 kinase The main objective of this project was to determine how these proteins are affecting the two actin driven particle uptake processes, phagocytosis and fluid uptake. We showed that ablation of PsrA gene which codes the regulatory subunit of PP2A resulted in a defective phagocytosis, whereas the fluid uptake was normal. We also showed for the first time that there was an increase in the phosphorylation of some of the PKB substrate proteins in wild type cell. Cells lacking PsrA gene displayed an aberrant phosphorylation of PKB substrate protein when compared to the wild type cells further confirming the involvement of PKB substrate in phagocytosis. Further, we looked at the effects of LKB1 kinase on phagocytosis by using a LKB1 knockdown construct introduced into wild type cells. The knock down of LKB1 resulted in a higher rate of phagocytosis while introduction of a LKB1 over expressing construct severally decreased the rate of phagocytosis indicating an inhibitory effect of LKB1. Furthermore there was an increase in the PKB substrate protein but a different pattern compared to the psrA- cells. We also carried out adhesion assays on LKB1 knockdown cells and the results showed a higher substrate adhesion as compared to the wild type cells, while psrA- cells had no adhesion defect.

Dictyostelium discoideum is a simple model widely used to study many cellular functions, including differentiation, gene regulation, cellular trafficking and directional migration. Adaptation mechanisms are essential in the regulation of these cellular processes. The misregulation of adaptation components often results in persistent activation of signaling pathways and aberrant cellular responses. Studying adaptation mechanisms regulating cellular migration will be crucial in the treatment of many pathological conditions in which motility plays a central role, such as tumor metastasis and acute inflammation. I will describe two adaptation mechanisms regulating directional migration in Dictyostelium cells. The Extracellular signal Regulated Kinase 2 (ERK2) plays an essential role in Dictyostelium cellular migration. ERK2 stimulates intracellular cAMP accumulation in chemotaxing cells. Aberrant ERK2 regulation results in aberrant cAMP levels and defective directional migration. The MAP Phosphatase with Leucine-rich repeats (MPL1) is crucial for ERK2 adaptation. Cells lacking, MPL1 (mpl1- cells) displayed higher pre-stimulus and persistent post-stimulus ERK2 phosphorylation, defective cAMP production and reduced cellular migration. Reintroduction of a full length Mpl1 into mpl1- cells restored aggregation, ERK2 regulation, random and directional motility, and cAMP production similar to wild type cells (Wt). These results suggest Mpl1 is essential for proper regulation of ERK2 phosphorylation and optimal motility in Dictyostelium cells. Cellular polarization in Dictyostelium cells in part is regulated by the activation of the AGC-related kinase Protein Kinase Related B1 (PKBR1). The PP2A regulatory subunit, B56, and the Glycogen Synthase Kinase 3 (GSK3) are necessary for PKBR1 adaptation in Dictyostelium cells. Cells lacking B56, psrA-cells, exhibited high basal and post-stimulus persistent phosphorylation of PKBR1, increased phosphorylation of PKBR1 substrates, and aberrant motility. PKBR1 adaptation is also regulated by the GSK3. When the levels of active GSK3 are reduced in Wt and psrA- cells, high basal levels of phosphorylated PKBR1 were observed, in a Ras dependent, but B56 independent mechanism. Altogether, PKBR1 adaptation is regulated by at least two independent mechanisms: one by GSK3 and another by PP2A/B56.

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