Loss of MYO5B in Mice Recapitulates Microvillus Inclusion Disease and Reveals an Apical Trafficking Pathway Distinct to Neonatal Duodenum

Abstract

Background and Aims Inactivating mutations in MYO5B cause severe neonatal diarrhea in Microvillus Inclusion Disease. Loss of active MYO5B causes the formation of pathognomonic inclusions and aberrations in brush border enzymes. Methods We developed three mouse models of germline, constitutively intestinal targeted and inducible intestinal targeted deletion of MYO5B. The mice were evaluated for enterocyte cellular morphology. Results Germline MYO5B KO mice showed early diarrhea and failure to thrive with evident microvillus inclusions and loss of apical transporters in the duodenum. IgG was present within inclusions. Apical transporters were lost and inclusions were present in the duodenum, but were nearly absent in the ileum. VillinCre;MYO5BF/F mice showed similar pathology and morphological changes in duodenal enterocytes. In contrast, when MYO5B KO was induced with tamoxifen treatment at 8 weeks of age, VillinCreERT2;MYO5BF/F mice developed severe diarrhea with loss of duodenal brush border enzymes, but few inclusions were observed in enterocytes. However, if tamoxifen is administered to 2-day-old VillinCreERT2;MYO5BF/F mice, prominent microvillus inclusions were observed. Conclusions The microvillus inclusions that develop after MYO5B loss reveal the presence of an unrecognized apical membrane trafficking pathway in neonatal duodenal enterocytes. However, the diarrheal pathology after MYO5B loss is due to deficits in transporter presentation at the apical membrane in duodenal enterocytes.

The zebrafish goosepimples/myosin Vb mutant exhibits cellular attributes of human microvillus inclusion disease

“…Analysis of intestinal cells of MVID patients has shown the presence of large vesicular structures called inclusion bodies with microvillar components trapped within (Ameen and Salas, 2000, Reinshagen et al, 2002, Thoeni et al, 2014, van der Velde et al, 2013). Enterocytes in Rab8a, Myo5b knockout mice and Myo5b deficient Caco2 cells exhibit the presence of actin rings marked by phalloidin, a classic feature of MVID (Carton-Garcia et al, 2015, Ruemmele et al, 2010, Sato et al, 2007, Schneeberger et al, 2015, Weis et al, 2016). To check if such inclusion bodies are present in the enterocytes of gsp/myoVb mutant larvae, intestines of 6dpf larvae were stained with phalloidin.…”

“…So far, the knockdown of myosin Vb using si-RNA in the Caco2 cell line has been used as a model for MVID (Ruemmele et al, 2010, Thoeni et al, 2014). Very recently, Myo5b knockout mouse models exhibiting attributes of MVID have been published (Carton-Garcia et al, 2015, Schneeberger et al, 2015, Weis et al, 2016). Other animal models for this disease include Rab8, Rab11a and Cdc42 knockout mice, which also exhibit microvillus inclusions in the enterocytes (Sakamori et al, 2012, Sato et al, 2007, Sobajima et al, 2014).…”

The mucosal barrier at a glance

“…Together, these structures form the apical junctional complex, which is also home to proteins that direct epithelial polarization (Anderson et al, 2004). Polarization is essential to epithelial function and involves proper sorting and delivery of both newly synthesized proteins and those trafficking through intracellular compartments to the apical or basolateral plasma membrane (Weis et al, 2016;Yeaman et al, 1999). This depends, in part, on binding of the exocyst complex to the apical junctional complex through cell-adhesion proteins (Yeaman et al, 2004) in order to direct basolateral exocytosis (Grindstaff et al, 1998).…”

ZO-1 interactions with F-actin and occludin direct epithelial polarization and single lumen specification in 3D culture

“…This is followed by the formation of a pre-apical patch (PAP) that is demarcated by tight junctions and, through fluid accumulation and membrane repulsion, expansion to form a lumen (Bagnat et al, 2007;Bryant et al, 2010). Genesis of a single lumen requires coordination of polarized cell division Jaffe et al, 2008;Qin et al, 2010), fluid accumulation (Bagnat et al, 2007), membrane delivery (Weis et al, 2016) and a balance between the kinetics of proliferation, apoptosis and polarization (Cerruti et al, 2013;Martín-Belmonte et al, 2008). These cellular processes are highly regulated and do not occur in isolation.…”