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Title page for ETD etd-03262018-125428


Type of Document Dissertation
Author Hebron, Katie Elizabeth
Author's Email Address katie.hebron@gmail.com
URN etd-03262018-125428
Title ALCAM Dynamically Regulates Tumor Cell Adhesion Through Differential Proteolysis and a Novel Binding Partner, Tetraspanin CD151
Degree PhD
Department Cancer Biology
Advisory Committee
Advisor Name Title
Barbara Fingleton Committee Chair
Andries Zijlstra Committee Member
Deborah Lannigan Committee Member
Simon Hayward Committee Member
Keywords
  • cancer
  • metastasis
  • bladder cancer
  • protein processing
  • alternative splicing
Date of Defense 2018-01-12
Availability restricted
Abstract
Metastasis persists as a significant unsolved hurdle in cancer treatment, with greater than 90% of cancer-related deaths attributed to metastasis. In order for cells to successfully metastasize, they must dynamically regulate cell adhesion. However, cell adhesion molecules are rarely mutated or deleted genetically in cancer, indicating that tumor cells are able to co-opt intrinsic regulatory mechanisms of adhesion to drive metastasis. We previously identified Activated Leukocyte Cell Adhesion Molecule (ALCAM) as a clinically relevant driver of metastasis and hypothesized that tunable regulation of its function contributes to tumor cell adhesion and metastasis. We tested this hypothesis through two channels.

We identified ALCAM as a novel binding partner of tetraspanin CD151, a known regulator of cell adhesion and motility. We previously demonstrated that clustering of integrin-free CD151 (CD151free) increased cell adhesion, decreased cell motility, and inhibited metastasis. Here, we identified ALCAM as a novel CD151 partner required for CD151free to control adhesion. Biochemical analyses revealed that CD151free is coupled to ALCAM by the scaffolding protein syntenin-1. Additionally, we show that the intracellular domain of ALCAM (ALCAM-ICD) is susceptible to ɣ-secretase cleavage, which releases a PDZ-binding peptide capable of disrupting the CD151/syntenin-1/ALCAM complex. Disruption of this complex impedes CD151free-mediated regulation of tumor cell adhesion and metastasis, demonstrating that CD151free controls tumor cell migration through a trimeric complex of CD151/syntenin-1/ALCAM.

Further evaluate of ALCAM revealed a potential alternative splicing which we predicted to control proteolytic shedding of its extracellular domain. We demonstrate that the loss of the membrane-proximal exon13 generates an ALCAM splice variant (ALCAM-Iso2) that enhances metastasis four-fold. Mechanistic studies identified a novel MMP14-dependent, membrane distal cleavage site in ALCAM-Iso2, which increases shedding ten-fold, thereby decreasing cellular cohesion and promoting motility. ALCAM-Iso2-expression was greatly increased in bladder cancer, further emphasizing that ALCAM alternative splicing can contribute to clinical disease progression. The requirement for both the loss of exon 13 and the gain of metalloprotease activity suggests that ALCAM shedding and concomitant regulation of dissemination is a locally tunable process.

In summary, this dissertation presents two mechanisms by which tumor cells are able to dynamically regulate cell adhesion to modulate migration and metastasis.

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