The role of receptor subunit interactions in formation and function of the high affinity interleukin-2 receptor

Date of Award




Degree Name

Doctor of Philosophy (Ph.D.)


Microbiology and Immunology

First Committee Member

Thomas R. Malek - Committee Chair


To assess the relative functional contribution of the $\beta$ and $\gamma\sb{\rm c}$ subunits of the interleukin 2 receptor (IL-2R), chimeric receptors, composed of the extracellular and transmembrane domains of human CD8$\alpha$ and the cytoplasmic domains of IL-2R $\beta$ and $\gamma\sb{\rm c}$, were generated and expressed in IL-2 dependent CTLL cells that undergo apoptosis upon IL-2 deprivation. CTLL that co-expressed CD8$\alpha$/IL-2R$\beta$ and CD8$\alpha$/IL-2R$\gamma$ were resistant to apoptosis and exhibited growth factor independent proliferation. This result demonstrates that stable heterodimerization of the $\beta$ and $\gamma\sb{\rm c}$ cytoplasmic domains is sufficient to produce a transformed phenotype in these T cells. By contrast, CTLL that expressed only CD8$\alpha$/IL-2R$\beta$ or CD8$\alpha$/IL-2R$\gamma$ were strictly dependent upon IL-2 for growth. CTLL that expressed CD8$\alpha$/IL-2R$\beta$ chimeric molecules were distinguished from parental CTLL by exhibiting a reduced rate of apoptosis after withdrawal of IL-2. After expression of CD8$\alpha$/IL-2R$\gamma\sb{c}$ chimeric molecules, several changes were noted for CTLL including an irregular morphology, a majority of cells that contained polyploid DNA, a reduced growth rate to IL-2, and a more rapid entry into apoptosis upon IL-2 withdrawal. These findings suggest that the IL-2R$\beta$ and $\gamma\sb{\rm c}$ subunits may be linked to independent signal transduction pathways that regulate apoptosis, but by themselves are incapable of inducing cell growth.To address the contribution of individual subunits in stabilizing the high affinity IL-2R, cell lines were produced that express hybrid human/mouse IL-2R composed of heterologous $\alpha$ and $\beta$ chains. Quantitative ligand binding studies and FACS analysis were performed for ELA cells that express hybrid human/mouse IL2R, and these data were compared to cells expressing homologous mouse IL-2R subunits. ELA cells that expressed human $\beta$/mouse $\alpha\gamma\sb{\rm c}$ formed high affinity IL-2R comparable to cells that expressed homologous mouse $\alpha$ and $\beta$ subunits. By contrast, EL4 cells that expressed homologous mouse IL-2R contained an approximately 10-fold higher level of high affinity IL-2R than EL4 cells that expressed heterologous human $\alpha$/mouse $\beta\gamma\sb{\rm c}$ IL-2R. This difference was not accounted for by lower levels of human $\alpha$ chains in these cell lines or lower expression of the mouse IL-2R$\gamma\sb{c}$ subunit as assessed by Northern blot analysis. Thus, these results suggest that amino acid sequences within the $\alpha$ subunit, distinct from the IL-2 binding site, are important in the assembly and stabilization of the high affinity IL-2R.These data suggest at least two roles for the IL2-R$\alpha$ subunit in formation of the high affinity IL-2R; one in ligand binding, the other in a molecular interaction with the IL-2R$\beta$ and/or IL-2R$\gamma\sb{c}$ subunits. However, IL-2 binding and internalization of ligand/receptor complexes are not necessary for IL-2R signal transduction, as stable dimerization of the IL-2R$\beta$ and IL-2R$\gamma\sb{c}$ subunits cytoplasmic domains alone was sufficient to induce the proliferation of CTLL cells. Thus, formation of a functional high affinity IL-2/IL-2R complex is the result of both ligand binding and interactions between sequences within the extracellular and cytoplasmic domains of the individual IL-2R subunits. (Abstract shortened by UMI.)


Biology, Molecular

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