TMSB4Y is a Candidate Tumor Suppressor on the Y Chromosome and is Deleted in Male Breast Cancer.

Male breast cancer comprises less than 1% of breast cancer diagnoses. Although estrogen exposure has been causally linked to the development of female breast cancers, the etiology of male breast cancer is unclear. Here, we show via fluorescence in situ hybridization (FISH) and droplet digital PCR (ddPCR) that the Y chromosome was clonally lost at a frequency of ~16% (5/31) in two independent cohorts of male breast cancer patients. We also show somatic loss of the Y chromosome gene TMSB4Y in a male breast tumor, confirming prior reports of loss at this locus in male breast cancers. To further understand the function of TMSB4Y, we created inducible cell lines of TMSB4Y in the female human breast epithelial cell line MCF-10A. Expression of TMSB4Y resulted in aberrant cellular morphology and reduced cell proliferation, with a corresponding reduction in the fraction of metaphase cells. We further show that TMSB4Y interacts directly with β-actin, the main component of the actin cytoskeleton and a cell cycle modulator. Taken together, our results suggest that clonal loss of the Y chromosome may contribute to male breast carcinogenesis, and that the TMSB4Y gene has tumor suppressor properties

Electron Tomography of the Contact between T Cells and SIV/HIV-1: Implications for Viral Entry

The envelope glycoproteins of primate lentiviruses, including human and simian immunodeficiency viruses (HIV and SIV), are heterodimers of a transmembrane glycoprotein (usually gp41), and a surface glycoprotein (gp120), which binds CD4 on target cells to initiate viral entry. We have used electron tomography to determine the three-dimensional architectures of purified SIV virions in isolation and in contact with CD4+ target cells. The trimeric viral envelope glycoprotein surface spikes are heterogeneous in appearance and typically ∼120 Å long and ∼120 Å wide at the distal end. Docking of SIV or HIV-1 on the T cell surface occurs via a neck-shaped contact region that is ∼400 Å wide and consistently consists of a closely spaced cluster of five to seven rod-shaped features, each ∼100 Å long and ∼100 Å wide. This distinctive structure is not observed when viruses are incubated with T lymphocytes in the presence of anti-CD4 antibodies, the CCR5 antagonist TAK779, or the peptide entry inhibitor SIVmac251 C34. For virions bound to cells, few trimers were observed away from this cluster at the virion–cell interface, even in cases where virus preparations showing as many as 70 envelope glycoprotein trimers per virus particle were used. This contact zone, which we term the “entry claw”, provides a spatial context to understand the molecular mechanisms of viral entry. Determination of the molecular composition and structure of the entry claw may facilitate the identification of improved drugs for the inhibition of HIV-1 entry

Hotspot SF3B1 mutations induce metabolic reprogramming and vulnerability to serine deprivation.

Cancer-associated mutations in the spliceosome gene SF3B1 create a neomorphic protein that produces aberrant mRNA splicing in hundreds of genes, but the ensuing biologic and therapeutic consequences of this missplicing are not well understood. Here we have provided evidence that aberrant splicing by mutant SF3B1 altered the transcriptome, proteome, and metabolome of human cells, leading to missplicing-associated downregulation of metabolic genes, decreased mitochondrial respiration, and suppression of the serine synthesis pathway. We also found that mutant SF3B1 induces vulnerability to deprivation of the nonessential amino acid serine, which was mediated by missplicing-associated downregulation of the serine synthesis pathway enzyme PHGDH. This vulnerability was manifest both in vitro and in vivo, as dietary restriction of serine and glycine in mice was able to inhibit the growth of SF3B1MUT xenografts. These findings describe a role for SF3B1 mutations in altered energy metabolism, and they offer a new therapeutic strategy against SF3B1MUT cancers

Phenotypic and Functional Properties of Helios+ Regulatory T Cells

Helios, an Ikaros family transcription factor, is preferentially expressed at the mRNA and protein level in regulatory T cells. Helios expression previously appeared to be restricted to thymic-derived Treg. Consistent with recent data, we show here that Helios expression is inducible in vitro under certain conditions. To understand phenotypic and functional differences between Helios+ and Helios− Treg, we profiled cell-surface markers of FoxP3+ Treg using unmanipulated splenocytes. We found that CD103 and GITR are expressed at high levels on a subset of Helios+ Treg and that a Helios+ Treg population could be significantly enriched by FACS sorting using these two markers. Quantitative real-time PCR (qPCR) analysis revealed increased TGF-β message in Helios+ Treg, consistent with the possibility that this population possesses enhanced regulatory potential. In tumor-bearing mice, we found that Helios+ Treg were relatively over-represented in the tumor-mass, and BrdU studies showed that, in vivo, Helios+ Treg proliferated more than Helios− Treg. We hypothesized that Helios-enriched Treg might exert increased suppressive effects. Using in vitro suppression assays, we show that Treg function correlates with the absolute number of Helios+ cells in culture. Taken together, these data show that Helios+ Treg represent a functional subset with associated CD103 and GITR expression

Analysis of Purified SIV Virions by Cryo-Electron Tomography

<div><p>(A) Low-dose projection image of a plunge-frozen specimen.</p><p>(B) A series of images recorded over a tilt range of −63° to +63° was used to reconstruct a 3-D volume of vitrified viruses similar to those shown in (A). Four 1-nm-thick tomographic slices at different depths from a tomogram of plunge-frozen SIV virions are shown.</p><p>Scale bars are 50 nm long. The dark black spots are from 5-nm-wide gold fiducial markers used for aligning individual images in a tilt series.</p></div

Electron Tomography of Virus–Cell Contact in Chronically and Acutely Infected Cells

<div><p>(A–C) Single 1-nm slices extracted from a dual axis tomogram reconstructed by weighted back-projection where part of an entry claw is captured in a chronically infected cell. One combination of three rods can be seen in one plane (A), while a different combination of three rods can be seen in the other (B). Four of the rods can be seen clearly in orthogonal view sectioned close to the plane of contact between virus and cell (C), with the densities arranged in a zig-zag manner. The black arrows point to the four densities in both transverse and top views.</p><p>(D) A 1-nm tomographic slice from SIV–T cell contact region in cells fixed 15 min after warming to 37 °C following incubation with high viral concentrations.</p><p>Scale bars are 100 nm long in (A–D).</p><p>(E) 3-D rendering of the same contact region presented in panel (D), showing the viral envelope (magenta), contact rods (red), core (yellow), and cell membrane (blue). Note that there are almost no spikes on the virion surface away from the region of viral–cell contact.</p></div

Analysis of Virus–Cell Contact under Different Conditions

<div><p>(A) Projection image of entry claw formed between SIV mac239 wild-type (full-length tail) and T cells fixed after incubation at 37 °C for 3 h.</p><p>(B) Projection image recorded from samples where SIV mac239 tail-truncated virus was fixed without warming to 37 °C. While viruses could be occasionally found in close proximity of the cell membrane, no entry claw–like structures were observed.</p><p>(C and D) Projection images recorded from cells incubated with SIV mac239 tail-truncated virus in the presence of 10 μM TAK779 or 5 μM C34 peptide, respectively. In both cases the viruses were incubated with cells for 3 h at 37 °C, and no entry claw–like structures could be detected.</p><p>Scale bars are 100 nm in all panels.</p></div

Projection Images from SIV-Infected Cells and from T Cells Exposed to SIV for Short Periods

<div><p>(A–C) Electron microscopic images of budding (A), immature (B), and mature (C) SIV particles in chronically infected cell suspensions. Scale bars are 50 nm long.</p><p>(D and E) Distinct virus–cell contacts in chronically infected cells that are defined by a characteristic density at the interface between the virus and the cell membrane, and distinct from virus morphologies seen in (A–C). Note that in some instances (E), the curvature of the cell membrane follows the curvature of the virus where the contact is made. Scale bars are 100 nm long.</p><p>(F) Projection image (higher magnifications shown in inset) of the contact region between SIV virions and T lymphocytes fixed after incubation at 37 °C for 15 min. At lower magnifications the overall context of the cell in the vicinity of the contact region is shown, while at the higher magnifications, entry claw contacts can be recognized in the projection view. Scale bar is 1 μm long. G, part of the Golgi ribbon; m, mitochondria; n, nucleus.</p></div