A Na,K-ATPase-Fodrin-Actin Membrane Cytoskeleton Complex is Required for Endothelial Fenestra Biogenesis.

Meihua Ju,Eija Jokitalo,Olli Rämö,Sofia Ioannidou,Helena Vihinen,David T. Shima,Peter Munro

doi:10.3390/cells9061387

Meihua Ju, Eija Jokitalo + Show 5 more

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https://doi.org/10.3390/cells9061387

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Abstract

Fenestrae are transcellular plasma membrane pores that mediate blood–tissue exchange in specialised vascular endothelia. The composition and biogenesis of the fenestra remain enigmatic. We isolated and characterised the protein composition of large patches of fenestrated plasma membrane, termed sieve plates. Loss-of-function experiments demonstrated that two components of the sieve plate, moesin and annexin II, were positive and negative regulators of fenestra formation, respectively. Biochemical analyses showed that moesin is involved in the formation of an actin–fodrin submembrane cytoskeleton that was essential for fenestra formation. The link between the fodrin cytoskeleton and the plasma membrane involved the fenestral pore protein PV-1 and Na,K-ATPase, which is a key regulator of signalling during fenestra formation both in vitro and in vivo. These findings provide a conceptual framework for fenestra biogenesis, linking the dynamic changes in plasma membrane remodelling to the formation of a submembrane cytoskeletal signalling complex.

Highlights

The precise regulation of blood–tissue interchange is critical for integration of the microvasculature and virtually all organ systems
Antibodies to moesin, phosphorylated moesin, and ezrin were from Cell Signalling (London, UK); antibody to annexin II was from Santa Cruz (Dallas, TX, USA); antibodies to β-tubulin and β-actin were from Sigma; antibodies to CD31 and fodrin were from Abcam (Cambridge, UK); and antibody to Na,K-ATPase was from Millipore (Feltham, UK)
The assembly of this fodrin cytoskeleton complex is promoted by the action of the plasma membrane–actin linker protein moesin and negatively regulated by the action of the pleiotropic actin-binding protein annexin II

Summary

Introduction

The precise regulation of blood–tissue interchange is critical for integration of the microvasculature and virtually all organ systems. Endothelial cells adopt highly specialised features to mediate the exchange of fluids and macromolecules across the vascular wall [1]. The blood–brain barrier consists of fortified cellular junctions which strictly restrict access of macromolecules to neural tissue [2]. At the other extreme are endothelial fenestrae, transcellular plasma membrane pores that passively mediate bidirectional blood–tissue exchange in the microvasculature of numerous organs, primarily with endocrine, absorptive, or filtrating functions [3]. Fenestrae are assumed to be central to many essential functions of the vasculature, such as the production of primary urine from blood [4], the sieving of lipoprotein particles [5], and the rapid vascular access and systemic dissemination of hormones from endocrine organs [6], though little direct experimentation exists.

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