Endocytosis of tight junction proteins and the regulation of degradation and recycling

Abstract

Internalization of tight junction (TJ) proteins from the plasma membrane is a pivotal mechanism regulating TJ plasticity and function in both epithelial and endothelial barrier tissues. Once internalized, the TJ proteins enter complex vesicular machinery, where further trafficking is directly dependent on the initiating stimulus and downstream signaling pathways that regulate the sorting and destiny of TJ proteins, as well as on cell and barrier responses. The destiny of internalized TJ proteins is recycling to the plasma membrane or sorting to late endosomes and degradation. This review highlights recent advances in our knowledge of endocytosis and vesicular trafficking of TJ proteins in both epithelial and endothelial cells. A greater understanding of these processes may allow for the development of methods to modulate barrier permeability for drug delivery or prevent barrier dysfunction in disease states.

Keywords: claudin; degradation; early endosomes; late endosomes; recycling.

Figure 1

Endocytic sorting of tight junction (TJ) proteins. Those proteins may be internalized by different mechanisms: clathrin-, caveolae-, and flotillin-dependent endocytosis or macropinocytosis. The TJ protein (cargo) is then sorted in early endosomes (Rab5⁺, EEA1⁺) and further directed toward either of two pathways: recycling (fast or slow) or degradation.

Figure 2

Rab5A and Rab5C regulate claudin-5 sorting. In mouse brain endothelial cells, CCL2 treatment (10 ng/mL) induced caveolae-dependent internalization of claudin-5 and its sorting toward Rab5⁺/EEA1⁺ early endosomes. (A) In parallel, CCL2 increased activation of Rab5 isoforms Rab5A and Rab5C, measured by Rab5A and Rab5C activation assays (Abcam, MA). The claudin-5 sorting process in CCL2-treated cells is directed from early endosomes primarily toward recycling (Rab4⁺), with a small portion in late (Rab7⁺) endosomes. (B) Analysis of the recycling and late endosomes–isolated fraction showed that, in condition of knockdown of Rab5A (Rab5A siRNA), claudin-5 sorting shifted from Rab7⁺ endosomes to recycling (Rab4⁺) endosomes. In contrast, depletion of Rab5C (Rab5C siRNA) caused accumulation of claudin-5 in Rab7⁺ endosomes. Overexpression of Rab5A and Rab5C induced increased accumulation in Rab7⁺ and Rab4⁺ endosomes, respectively. These data indicate the involvement of Rab5A and Rab5C in claudin-5 sorting, with predominant sorting of Rab5a to late endosomes and degradation and Rab5C to a recycling pathway. IP, immmunoprecipitation.