The intestinal epithelium is the most rapidly self-renewing tissue in adult mammals. We have recently demonstrated the presence of about six cycling Lgr5(+) stem cells at the bottoms of small-intestinal crypts. Here we describe the establishment of long-term culture conditions under which single crypts undergo multiple crypt fission events, while simultanously generating villus-like epithelial domains in which all differentiated cell types are present. Single sorted Lgr5(+) stem cells can also initiate these cryptvillus organoids. Tracing experiments indicate that the Lgr5(+) stem-cell hierarchy is maintained in organoids. We conclude that intestinal cryptvillus units are self-organizing structures, which can be built from a single stem cell in the absence of a non-epithelial cellular niche.

Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche.

Collagen is the most abundant protein in animals. This fibrous, structural protein comprises a right-handed bundle of three parallel, left-handed polyproline II-type helices. Much progress has been made in elucidating the structure of collagen triple helices and the physicochemical basis for their stability. New evidence demonstrates that stereoelectronic effects and preorganization play a key role in that stability. The fibrillar structure of type I collagen—the prototypical collagen fibril—has been revealed in detail. Artificial collagen fibrils that display some properties of natural collagen fibrils are now accessible using chemical synthesis and self-assembly. A rapidly emerging understanding of the mechanical and structural properties of native collagen fibrils will guide further development of artificial collagenous materials for biomedicine and nanotechnology.

/pdf/collagen-structure-and-stability-4a8qztbno6.pdf

Collagen Structure and Stability

https://www.gastrojournal.org/article/S0016-5085(11)01108-5/pdf

Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett's epithelium.

Publisher Summary This chapter discusses the role of plasminogen activators in various biological processes. In specific, it describes two types of plasminogen activators—namely, the urokinase-type plasminogen activator (u-PA) and the tissue-type plasminogen activator (t-PA), which are essentially different gene products. The amino acid sequences of these activators and nucleotide sequences of the corresponding cDNAs have largely been determined, and the cDNAs have been cloned using recombinant techniques. A variety of enzymatic as well as immunological assay and detection methods have also been developed that allows a precise quantification of the activators, a distinction between u-PA and t-PA, determination of whether an activator is present in its active or zymogen form, analysis of the kinetics of different steps of the cascade reaction, and immunocytochemical identification of u-PA and t-PA in tissue sections. Much of the studies on plasminogen activators and cancer has been guided by the hypothesis that proteolysis of the components of extracellular matrix, initiated by the release of plasminogen activator from the cancer cells, plays a decisive role for the degradation of normal tissue, and thereby for invasive growth and metastases.

Plasminogen activators, tissue degradation, and cancer.

Prions are composed largely, if not entirely, of prion protein (PrPSc in the case of scrapie). Although the formation of PrPSc from the cellular prion protein (PrPC) is a post-translational process, no candidate chemical modification was identified, suggesting that a conformational change features in PrPSc synthesis. To assess this possibility, we purified both PrPC and PrPSc by using nondenaturing procedures and determined the secondary structure of each. Fourier-transform infrared (FTIR) spectroscopy demonstrated that PrPC has a high alpha-helix content (42%) and no beta-sheet (3%), findings that were confirmed by circular dichroism measurements. In contrast, the beta-sheet content of PrPSc was 43% and the alpha-helix 30% as measured by FTIR. As determined in earlier studies, N-terminally truncated PrPSc derived by limited proteolysis, designated PrP 27-30, has an even higher beta-sheet content (54%) and a lower alpha-helix content (21%). Neither PrPC nor PrPSc formed aggregates detectable by electron microscopy, while PrP 27-30 polymerized into rod-shaped amyloids. While the foregoing findings argue that the conversion of alpha-helices into beta-sheets underlies the formation of PrPSc, we cannot eliminate the possibility that an undetected chemical modification of a small fraction of PrPSc initiates this process. Since PrPSc seems to be the only component of the "infectious" prion particle, it is likely that this conformational transition is a fundamental event in the propagation of prions.

https://www.pnas.org/content/pnas/90/23/10962.full.pdf

Conversion of alpha-helices into beta-sheets features in the formation of the scrapie prion proteins.

http://www.jbc.org/content/254/19/9933.full.pdf

Laminin--a glycoprotein from basement membranes.

Collagen type IV, laminin, heparan sulfate proteoglycans, nidogen (entactin) and BM-40 (osteonectin, SPARC) represent major structural proteins of basement membranes. They are well-characterized in their domain structures, amino acid sequences and potentials for molecular interactions. Such interactions include self-assembly processes and heterotypic binding between individual constituents, as well as binding of calcium (laminin, BM-40) and are likely to be used for basement membrane assembly. Laminin, collagen IV and nidogen also possess several cell-binding sites which interact with distinct cellular receptors. Some evidence exists that those interactions are involved in the control of cell behaviour. These observations have provided a more defined understanding of basement membrane function and the definition of new research goals in the future.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1989.tb14673.x

Structure and biological activity of basement membrane proteins

CONTENTS INTRODUCTION.. .• .••.....• .•••.. 57 STRUCTURE 58 INTERACTIONS WITH OTHER BASEMENT MEMBRANE PROTEINS ......•• 65 BIOSYNTHESIS AND REGULATION •.•....• • • 69 CELL BINDING AND OTHER BIOLOGICAL PROPERTIES OF LAMININ 71 MOLECULAR MODELS OF BASEMENT MEMBRANE STRUCTURE •.. 76

Laminin and Other Basement Membrane Components

A new nomenclature for the laminins.

Basement membranes are thin sheets of extracellular proteins situated in close contact with cells at various locations in the body. They have a great influence on tissue compartmentalization and cellular phenotypes from early embryonic development onwards. The major constituents of all basement membranes are collagen IV and laminin, which both exist as multiple isoforms and each form a huge irregular network by self assembly. These networks are connected by nidogen, which also binds to several other components (proteoglycans, fibulins). Basement membranes are connected to cells by several receptors of the integrin family, which bind preferentially to laminins and collagen IV, and via some lectin-type interactions. The formation of basement membranes requires cooperation between different cell types since nidogen, for example, is usually synthesized by cells other than those exposed to the basement membranes. Thus many molecular interactions, of variable affinities, determine the final shape of basement membranes and their preferred subanatomical localization.

Rupert Timpl

Papers

Laminin--a glycoprotein from basement membranes.

Structure and biological activity of basement membrane proteins

Laminin and Other Basement Membrane Components

A new nomenclature for the laminins.

Supramolecular assembly of basement membranes