AI-driven designed protein epigenomics.

The biological revolutions of computationally designed proteins, induced pluripotent stem cells (iPSCs), and the CRISPR-Cas9 system finally enables modifications that can spur deep understanding of spatial requirement of epigenetic information. This commentary describes the utility of a computationally designed protein, EED Binder (EB), when fused to dCas9 (EBdCas9) for identifying critical sites of PRC2 dependent histone H3K27me3 marks in the chromatin. By using EBdCas9 and gRNA, PRC2 function can be inhibited at specific loci, resulting in precise reduction of EZH2 and H3K27me3 marks, and in some (but not all) locations, activation of the gene and functional outcomes (such as regulation of cell cycle or trophoblast transdifferentiation).

Prolactinoma: Medical and Surgical Considerations.

Prolactinomas are the most common secretory tumor of the pituitary gland. Clinical symptoms may be due to prolactin oversecretion, localized mass effect, or a combination of both. Although the mainstay of prolactinoma management is medical therapy with dopamine agonists, endoscopic endonasal or transcranial surgery, radiation therapy, or a combination of these is an important treatment option in select cases.

dCas9 fusion to computer-designed PRC2 inhibitor reveals functional TATA box in distal promoter region.

Bifurcation of cellular fates, a critical process in development, requires histone 3 lysine 27 methylation (H3K27me3) marks propagated by the polycomb repressive complex 2 (PRC2). However, precise chromatin loci of functional H3K27me3 marks are not yet known. Here, we identify critical PRC2 functional sites at high resolution.

Amino acid primed mTOR activity is essential for heart regeneration.

Heart disease is the leading cause of death with no method to repair damaged myocardium due to the limited proliferative capacity of adult cardiomyocytes. Curiously, mouse neonates and zebrafish can regenerate their hearts via cardiomyocyte de-differentiation and proliferation. However, a molecular mechanism of why these cardiomyocytes can re-enter cell cycle is poorly understood.

Author Correction: TFPa/HADHA is required for fatty acid beta-oxidation and cardiolipin re-modeling in human cardiomyocytes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Epigenetic metabolites license stem cell states.

It has become clear during recent years that stem cells undergo metabolic remodeling during their activation process. While these metabolic switches take place in pluripotency as well as adult stem cell populations, the rules that govern the switch are not clear. In this review, we summarize some of the transitions in adult and pluripotent cell types and will propose that the key function in this process is the generation of epigenetic metabolites that govern critical epigenetic modifications, and therefore stem cell states.

UTILITY OF AFIRMA GENE EXPRESSION CLASSIFIER FOR EVALUATION OF INDETERMINATE THYROID NODULES AND CORRELATION WITH ULTRASOUND RISK ASSESSMENT: SINGLE INSTITUTIONAL EXPERIENCE.

We assessed our experience with Afirma gene expression classifier (GEC) combined with sono-graphic risk assessment, using both the American Thyroid Association (ATA) and the Thyroid Imaging Reporting and Data System (TI-RADS) in evaluating indeterminate thyroid nodules. We identified 98 patients with 101 nodules who had a second fine needle aspiration biopsy (FNA) between January 1, 2014, and September 30, 2017, and sent to Veracyte for cytopathology and subsequent Afirma GEC testing. A second FNA biopsy was performed if the initial cytopathology was either Bethesda III or IV (n = 94) or nondiagnostic (n = 7).

Pituitary Dysfunction Among Men Presenting with Hypogonadism.

Purpose Of Review: Hypogonadism is a common endocrine dysfunction. This review focuses on the most up-to-date guideline for evaluation of pituitary function among men presenting with signs and symptoms of hypogonadism.

Recent Findings: The clinician must differentiate between primary (testicular) and secondary (pituitary-hypothalamic or central) hypogonadisms and be aware of adult-onset hypogonadism.

TFPa/HADHA is required for fatty acid beta-oxidation and cardiolipin re-modeling in human cardiomyocytes.

Mitochondrial trifunctional protein deficiency, due to mutations in hydratase subunit A (HADHA), results in sudden infant death syndrome with no cure. To reveal the disease etiology, we generated stem cell-derived cardiomyocytes from HADHA-deficient hiPSCs and accelerated their maturation via an engineered microRNA maturation cocktail that upregulated the epigenetic regulator, HOPX.  Here we report, matured HADHA mutant cardiomyocytes treated with an endogenous mixture of fatty acids manifest the disease phenotype: defective calcium dynamics and repolarization kinetics which results in a pro-arrhythmic state.

Single-Cell Transcriptomic Analysis of Cardiac Differentiation from Human PSCs Reveals HOPX-Dependent Cardiomyocyte Maturation.

Cardiac differentiation of human pluripotent stem cells (hPSCs) requires orchestration of dynamic gene regulatory networks during stepwise fate transitions but often generates immature cell types that do not fully recapitulate properties of their adult counterparts, suggesting incomplete activation of key transcriptional networks. We performed extensive single-cell transcriptomic analyses to map fate choices and gene expression programs during cardiac differentiation of hPSCs and identified strategies to improve in vitro cardiomyocyte differentiation. Utilizing genetic gain- and loss-of-function approaches, we found that hypertrophic signaling is not effectively activated during monolayer-based cardiac differentiation, thereby preventing expression of HOPX and its activation of downstream genes that govern late stages of cardiomyocyte maturation.

Inducible CRISPR genome editing platform in naive human embryonic stem cells reveals JARID2 function in self-renewal.

To easily edit the genome of naïve human embryonic stem cells (hESC), we introduced a dual cassette encoding an inducible Cas9 into the AAVS1 site of naïve hESC (iCas9). The iCas9 line retained karyotypic stability, expression of pluripotency markers, differentiation potential, and stability in 5iLA and EPS pluripotency conditions. The iCas9 line induced efficient homology-directed repair (HDR) and non-homologous end joining (NHEJ) based mutations through CRISPR-Cas9 system.

First critical repressive H3K27me3 marks in embryonic stem cells identified using designed protein inhibitor.

The polycomb repressive complex 2 (PRC2) histone methyltransferase plays a central role in epigenetic regulation in development and in cancer, and hence to interrogate its role in a specific developmental transition, methods are needed for disrupting function of the complex with high temporal and spatial precision. The catalytic and substrate recognition functions of PRC2 are coupled by binding of the N-terminal helix of the Ezh2 methylase to an extended groove on the EED trimethyl lysine binding subunit. Disrupting PRC2 function can in principle be achieved by blocking this single interaction, but there are few approaches for blocking specific protein-protein interactions in living cells and organisms.

Zinc transport and the inhibition of the L-type calcium channel are two separable functions of ZnT-1.

Traditionally, proteins are considered to perform a single role, be it as an enzyme, a channel, a transporter or as a structural scaffold. However, recent studies have described moonlighting proteins that perform distinct and independent functions; for example, TRPM7 is both an ion channel and a kinase. ZnT-1 is a member of the Carrier Diffusion Facilitator family that is expressed throughout the phylogenetic tree from bacteria to humans.

Polyadenylation and degradation of RNA in the mitochondria.

Mitochondria have their own gene expression machinery and the relative abundance of RNA products in these organelles in animals is mostly dictated by their rate of degradation. The molecular mechanisms regulating the differential accumulation of the transcripts in this organelle remain largely elusive. Here, we summarize the present knowledge of how RNA is degraded in human mitochondria and describe the coexistence of stable poly(A) tails and the nonabundant tails, which have been suggested to play a role in the RNA degradation process.

Identification of LACTB2, a metallo-β-lactamase protein, as a human mitochondrial endoribonuclease.

Post-transcriptional control of mitochondrial gene expression, including the processing and generation of mature transcripts as well as their degradation, is a key regulatory step in gene expression in human mitochondria. Consequently, identification of the proteins responsible for RNA processing and degradation in this organelle is of great importance. The metallo-β-lactamase (MBL) is a candidate protein family that includes ribo- and deoxyribonucleases.

ZnT-1 extrudes zinc from mammalian cells functioning as a Zn(2+)/H(+) exchanger.

ZnT-1 is a Cation Diffusion Facilitator (CDF) family protein, and is present throughout the phylogenetic tree from bacteria to humans. Since its original cloning in 1995, ZnT-1 has been considered to be the major Zn(2+) extruding transporter, based on its ability to protect cells against zinc toxicity. However, experimental evidence for ZnT-1 induced Zn(2+) extrusion was not convincing.

Selective modulation of cellular voltage-dependent calcium channels by hyperbaric pressure-a suggested HPNS partial mechanism.

Professional deep sea divers experience motor and cognitive impairment, known as High Pressure Neurological Syndrome (HPNS), when exposed to pressures of 100 msw (1.1 MPa) and above, considered to be the result of synaptic transmission alteration. Previous studies have indicated modulation of presynaptic Ca(2+) currents at high pressure.

Pressure-selective modulation of NMDA receptor subtypes may reflect 3D structural differences.

Professional deep-water divers exposed to high pressure (HP) above 1.1 MPa suffer from High Pressure Neurological Syndrome (HPNS), which is associated with CNS hyperexcitability. We have previously reported that HP augments N-methyl-D-aspartate receptor (NMDAR) synaptic responses, increases neuronal excitability, and potentially causes irreversible neuronal damage.

ZnT-1 enhances the activity and surface expression of T-type calcium channels through activation of Ras-ERK signaling.

Zinc transporter-1 (ZnT-1) is a putative zinc transporter that confers cellular resistance from zinc toxicity. In addition, ZnT-1 has important regulatory functions, including inhibition of L-type calcium channels and activation of Raf-1 kinase. Here we studied the effects of ZnT-1 on the expression and function of T-type calcium channels.

ZnT-1 protects HL-1 cells from simulated ischemia-reperfusion through activation of Ras-ERK signaling.

Activation of ERK signaling may promote cardioprotection from ischemia-reperfusion (I/R) injury. ZnT-1, a protein that confers resistance from zinc toxicity, was found to interact with Raf-1 kinase through its C-terminal domain, leading to downstream activation of ERK. In the present study, we evaluated the effects of ZnT-1 in cultured murine cardiomyocytes (HL-1 cells) that were exposed to simulated-I/R.

Distinct activities of several RNase J proteins in methanogenic archaea.

RNA degradation plays an important role in the control of gene expression in all domains of life, including Archaea. While analyzing RNA degradation in different archaea, we faced an interesting situation. The members of a group of methanogenic archaea, including Methanocaldococcus jannaschii, contain neither the archaeal exosome nor RNase II/R homologs.

SpRET: highly sensitive and reliable spectral measurement of absolute FRET efficiency.

Contemporary research aims to understand biological processes not only by identifying participating proteins, but also by characterizing the dynamics of their interactions. Because Förster's Resonance Energy Transfer (FRET) is invaluable for the latter undertaking, its usage is steadily increasing. However, FRET measurements are notoriously error-prone, especially when its inherent efficiency is low, a not uncommon situation.

Molecular basis for zinc transporter 1 action as an endogenous inhibitor of L-type calcium channels.

The L-type calcium channel (LTCC) has a variety of physiological roles that are critical for the proper function of many cell types and organs. Recently, a member of the zinc-regulating family of proteins, ZnT-1, was recognized as an endogenous inhibitor of the LTCC, but its mechanism of action has not been elucidated. In the present study, using two-electrode voltage clamp recordings in Xenopus oocytes, we demonstrate that ZnT-1-mediated inhibition of the LTCC critically depends on the presence of the LTCC regulatory beta-subunit.

Markedly low hemoglobin A1c in a patient with an unusual presentation of beta-thalassemia minor.

Objective: To describe very low hemoglobin A1c levels in a patient with type 2 diabetes mellitus and an unusual presentation of beta-thalassemia minor.

Methods: We present the clinical and laboratory findings of the study patient.

Results: A 64-year-old African American man with type 2 diabetes mellitus was referred to the endocrinology clinic with a hemoglobin A1c level of 1.

Unusual mid-shaft fractures during long-term bisphosphonate therapy.

Background: Bisphosphonates are the most commonly prescribed medications for the treatment of osteoporosis. Although existing evidence supports a good safety profile, there is concern that chronic administration of these agents could result in severe suppression of bone turnover with increased risk of nonvertebral fractures.

Objective: The objective of this study was to report the clinical presentation, selected bone histomorphometry and X-ray images of patients who developed mid-shaft long bone fractures during bisphosphonate therapy, six of whom had bone biopsy for histomorphometery.