Abstract
The teneurins are a family of four transmembrane proteins essential to intercellular adhesion processes, and are required for the development and maintenance of tissues. The Adhesion G protein-coupled receptor (GPCR) subclass latrophilins (ADGRL), or simply the latrophilins (LPHN), are putative receptors of the teneurins and act, in part, to mediate intercellular adhesion via binding with the teneurin extracellular region. At the distal tip of the extracellular region of each teneurin lies a peptide sequence termed the teneurin C-terminal associated peptide (TCAP). TCAP-1, associated with teneurin-1, is itself bioactive, suggesting that TCAP is a critical functional region of teneurin. However, the role of TCAP-1 has not been established with respect to its ability to interact with LPHN to induce downstream effects. To establish that TCAP-1 binds to LPHN1, a FLAG-tagged hormone binding domain (HBD) of LPHN1 and a GFP-tagged TCAP-1 peptide were co-expressed in HEK293 cells. Both immunoreactive epitopes were co-localized as a single band after immunoprecipitation, indicating an association between the two proteins. Moreover, fluorescent co-labeling occurred at the plasma membrane of LPHN1 over-expressing cells when treated with a FITC-tagged TCAP-1 variant. Expression of LPHN1 and treatment with TCAP-1 modulated the actin-based cytoskeleton in these cells in a manner consistent with previously reported actions of TCAP-1 and affected the overall morphology and aggregation of the cells. This study indicates that TCAP-1 may associate directly with LPHN1 and could play a role in the modulation of cytoskeletal organization and intercellular adhesion and aggregation via this interaction.
Highlights
The teneurins are a family of type II transmembrane proteins critical for the development and maintenance of the central nervous system in both vertebrates and invertebrates
It should be noted that endogenous expression of the LPHN2 and LPHN3 isoforms in HEK-WT and HEK-Puro cells (WT) and HEK-LPHN1-S cells was not examined in this study
The green fluorescence protein (GFP)-mTCAP1 construct, based on the putative 41-mer terminal associated peptide (TCAP)-1 region, was present as a band at approximately 30 kDa in the V444-E634 eluate and, to a lesser degree, in the V443-Q579 eluate (Figure 3, IPs, red arrow). This corresponds to the 30 kDa band observed for GFP-Mouse TCAP-1 (mTCAP-1) in the eluate inputs (Figure 3, inputs), and is consistent with a protein composed of the 25 kDa GFP and the 4.7 kDa mature TCAP-1 peptide. These results indicate that the mature TCAP-1 peptide may interact with LPHN1, most likely with a segment encompassing hormone binding domain (HBD) residues V444 to E634
Summary
The teneurins are a family of type II transmembrane proteins critical for the development and maintenance of the central nervous system in both vertebrates and invertebrates. Vertebrates contain four paralogous teneurins (teneurin-1 through -4), each of which are 2,500–2,800 residues in length and are comprised of numerous multifunctional domains involved in adhesion, cytoskeletal binding, and other protein-protein interactions [1,2,3,4,5] In both vertebrates and TCAP-1 and Latrophilin Modulate Adhesion invertebrates, the teneurins have been implicated in the formation of filopodia and outgrowth of neurites, as well as neuronal mapping, axonal path-finding, and increased cell-cell adhesion [6,7,8,9,10,11,12,13,14,15]. Despite the high efficacy TCAP-1 shows both in vitro and in vivo, the precise mechanism by which these actions occur is not well-understood
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