Dissemination of tumor cells is an essential step in metastasis. Direct contact between a macrophage, Mena over-expressing tumor cell and endothelial cell [Tumor MicroEnvironment of Metastasis (TMEM)], correlates with metastasis in breast cancer patients. Here we show, using intravital high-resolution two-photon microscopy, that transient vascular permeability and tumor cell intravasation occur simultaneously and exclusively at TMEM. The hyperpermeable nature of tumor vasculature is described as spatially and temporally heterogeneous. Using real-time imaging we observed that vascular permeability is transient, restricted to TMEM, and required for tumor cell dissemination. VEGFA signaling from Tie2Hi TMEM macrophages causes local loss of vascular junctions, transient vascular permeability and tumor cell intravasation, demonstrating a role for TMEM within the primary mammary tumor. These data provide insight into the mechanism of tumor cell intravasation and vascular permeability in breast cancer, explaining the value of TMEM density as a predictor of distant metastatic recurrence in patients.

Lampreys are representatives of an ancient vertebrate lineage that diverged from our own ~500 million years ago. By virtue of this deeply shared ancestry, the sea lamprey (P. marinus) genome is uniquely poised to provide insight into the ancestry of vertebrate genomes and the underlying principles of vertebrate biology. Here, we present the first lamprey whole-genome sequence and assembly. We note challenges faced owing to its high content of repetitive elements and GC bases, as well as the absence of broad-scale sequence information from closely related species. Analyses of the assembly indicate that two whole-genome duplications likely occurred before the divergence of ancestral lamprey and gnathostome lineages. Moreover, the results help define key evolutionary events within vertebrate lineages, including the origin of myelin-associated proteins and the development of appendages. The lamprey genome provides an important resource for reconstructing vertebrate origins and the evolutionary events that have shaped the genomes of extant organisms.

Monomeric near-infrared (NIR) fluorescent proteins (FPs) are in high demand as protein tags and components of biosensors for deep-tissue imaging and multicolour microscopy. We report three bright and spectrally distinct monomeric NIR FPs, termed miRFPs, engineered from bacterial phytochrome, which can be used as easily as GFP-like FPs. miRFPs are 2–5-fold brighter in mammalian cells than other monomeric NIR FPs and perform well in protein fusions, allowing multicolour structured illumination microscopy. miRFPs enable development of several types of NIR biosensors, such as for protein–protein interactions, RNA detection, signalling cascades and cell fate. We demonstrate this by engineering the monomeric fluorescence complementation reporters, the IκBα reporter for NF-κB pathway and the cell cycle biosensor for detection of proliferation status of cells in culture and in animals. miRFPs allow non-invasive visualization and detection of biological processes at different scales, from super-resolution microscopy to in vivo imaging, using the same probes.

Jawless vertebrates use variable lymphocyte receptors (VLR) comprised of leucine-rich-repeat (LRR) segments as counterparts of the immunoglobulin based receptors that jawed vertebrates use for antigen recognition. Highly diverse VLR genes are somatically assembled by the insertion of variable LRR sequences into incomplete germline VLRA and VLRB genes. Here we show that VLRA and VLRB anticipatory receptors are expressed by separate lymphocyte populations through monoallelic VLRA or VLRB assembly in concert with expression of Cytosine deaminase 1 or Cytosine deaminase 2, respectively. Distinctive gene expression profiles for VLRA+ and VLRB+ lymphocytes resemble those of mammalian T and B cells. Although both VLRA and VLRB cells proliferate in response to antigenic stimulation, only the VLRB lymphocytes bind native antigens and differentiate into VLR antibody secreting cells. Conversely, VLRA lymphocytes respond preferentially to a classical T cell mitogen and upregulate their expression of proinflammatory cytokine genes, IL-17 and MIF. The finding of T-like and B-like lymphocytes in lampreys offers new insight into the evolution of adaptive immunity.

Jawed vertebrates (gnathostomes) and jawless vertebrates (cyclostomes) have different adaptive immune systems1,2. Gnathostomes use T- and B-cell antigen receptors belonging to the immunoglobulin superfamily3,4. Cyclostomes, the lampreys and hagfish, instead use leucine-rich repeat proteins to construct variable lymphocyte receptors (VLRs), two types of which, VLRA and VLRB, are reciprocally expressed by lymphocytes resembling gnathostome T and B cells5–7. Here we define another lineage of T-cell-like lymphocytes that express the recently identified VLRC receptors8,9. Both VLRC+ and VLRA+ lymphocytes express orthologues of genes that gnathostome γδ and αβ T cells use for their differentiation, undergo VLRC and VLRA assembly and repertoire diversification in the ‘thymoid’ gill region, and express their VLRs solely as cell-surface proteins. Our findings suggest that the genetic programmes for two primordial T-cell lineages and a prototypic B-cell lineage were already present in the last common vertebrate ancestor approximately 500 million years ago. We propose that functional specialization of distinct T-cell-like lineages was an ancient feature of a primordial immune system.