Laminin N-terminus α31 expression during development is lethal and causes widespread tissue-specific defects in a transgenic mouse model

Abstract

Alternative splicing of the laminin α3 gene gives rise to a netrin-like protein termed LaNt α31, the major structural feature of which is a laminin N-terminal domain. LaNt α31 is expressed across a wide range of tissues, is upregulated in cancers, and ex vivo and in vitro functional studies have indicated that this relatively unstudied protein influences wound repair, stem cell activity, and tumour progression via modifying matrix organisation. However, LaNt α31 functionality has never been investigated in vivo. Here we report the generation and characterisation of the first LaNt α31 transgenic mouse line using the ubiquitin C promoter to drive expression of an expression cassette containing a flox-STOP sequence, the human LaNt α31 coding sequence and a tdTomato reporter. This line was crossed with mice expressing inducible Cre recombinase driven from the Rosa26 locus (R26CreERT2), and double transgenics were given tamoxifen at E15.5 to induce expression. LaNt α31 overexpressing animals were fully formed and intact at birth but were not viable, exhibiting localised regions of erythema. Histological examination revealed numerous striking defects includng extra-vascular erythrocytes across multiple tissues. Widespread disorganisation was apparent in the kidney, with epithelial detachment, tubular dilation, interstitial bleeding observed, and thickening of the kidney tubule basement membranes. In the skin, mice exhibited disruption of the epidermal basal cell layer and hair follicle outer root sheath, with evidence of basement membrane interruption in the interfollicular epidermis. In the liver, there was a ∼50% reduction of total cell number, associated with a depletion of hematopoietic erythrocytic foci. In the lungs, there appeared to be a reduction of alveolar epithelial cells accompanied by blood interspersed throughout the tissue. Together, these findings demonstrate that LaNt α31 can influence tissue morphogenesis during development and implicate this new protein as a potentially important mediator of basement membrane assembly.

Laminin Polymerization and Inherited Disease: Lessons From Genetics

“…One LAMA3-derived alternative splice isoform, Laminin N terminus α31 (LaNt α31) has widespread expression in human tissues (Troughton et al, 2020b), is upregulated during wounding and corneal limbal stem cell activation (Barrera et al, 2018) and emerging data indicate that it can modulate LM organization in vitro (Troughton et al, 2020a). In vivo overexpression is embryonic lethal during development with tissue defects that resemble LM network disruption phenotypes (Sugden et al, 2020).…”