bluuebunny/crossref_metadata_2025_split huggingface dataset:
IMP1 loss promotes enhanced recovery from acute colitis (899.3)
Authors: Hamilton Kathryn , Lundsmith Emma , Hahn Christopher , Noubissi Felicite , Spiegelman Vladimir , Rustgi Anil 
Published: June 2021

Background: mRNA‐binding proteins, including Mushashi1, Lin28b, and Igf2 mRNA‐binding protein 1 (IMP1), are newly recognized regulators of intestinal epithelial homeostasis via their effects on post‐transcriptional regulation of mRNAs comprising critical growth regulatory networks. Imp1‐null mice exhibit severe morphologic defects in the intestinal epithelium, yet the specific mechanisms of IMP1 in normal intestinal homeostasis and response to injury remain to be elucidated. We hypothesize that Imp1 modulates intestinal epithelial cell proliferation in response to acute inflammation.Methods: We generated mice with intestinal epithelial‐specific loss of Imp1 (VillinCre;Imp1flox/flox mice), treated with dextran sodium sulfate (DSS) and measured indices during recovery, as well as IL‐6 and IL‐1β, canonical pro‐inflammatory cytokines. Actively proliferating cells were evaluated using Ki67 staining.Results: Untreated VillinCre;Imp1floxed mice exhibit increased Ki67+ cells in the extra‐proliferative zone compared to controls. Upon DSS treatment followed by 2 days of recovery, VillinCre;Imp1flox/flox mice display decreased histological disease scores and lower cytokine expression compared to controls.Conclusions: VillinCre;Imp1flox/flox mice have enhanced recovery following DSS and a decreased immune response. Mislocalization of actively proliferating cells may be advantageous in this context, providing a novel link of IMP1 to intestinal epithelial regeneration.Grant Funding Source: Supported by: F32 DK093207‐01 (KEH), HHMI MRFP (ETL), NIH RO1 DK056645 (KEH, AKR), NIH UO1 DK085551
Ferristatin II induces hepatic hepcidin expression and Stat3 phosphorylation (907.3)
Authors: Alkhateeb Ahmed , Buckett Peter , Kim Jonghan , Byrne Shaina , Wessling‐Resnick Marianne 
Published: June 2021

Administration of Ferristatin II (FS2) to rats is associated with decreased serum iron, reduced transferrin saturation and increased hepatic hepcidin expression without changes in liver non‐heme iron levels. To better understand the mechanisms underlying hepcidin induction by FS2, its effects on ER stress were examined. XBP1 splicing, a marker of ER stress, was not altered in livers from treated rats. Likewise, levels of the ER stress protein Grp94 were unchanged in FS2‐treated Hep3B cells. SMAD signaling, a second regulatory pathway, did not appear to be activated since transcript levels of downstream targets SMAD7, BMP6 and Id1 were all reduced in livers of FS2‐treated rats. However, mRNA levels of alpha‐2‐microglobulin, alpha‐1‐acid glycoprotein and C‐reactive protein were up‐regulated. Western blot experiments confirmed that FS2 treatment induced phosphorylation of hepatic Stat3, a known mediator of the acute phase response and a regulator of hepcidin expression. Accordingly, FS2 induced Stat3 phosphorylation in HepG2 cells in vitro. These results suggest that FS2 induces hepatic hepcidin expression in a Stat3‐dependent manner. The pharmacological induction of hepcidin by ferristatin II holds clinical potential to decrease iron absorption in iron overload diseases.Grant Funding Source: Supported by NIH NIDDK (DK064750)
Inflammation reduces the activity of the NaHCO3 cotransporter, NBCe1, in the surface cells of the proximal colon of IL10 knockout mice (908.1)
Authors: Fan Shjun , Schultz Michael , Butt Grant 
Published: June 2021

The impact of inflammation on intestinal secretion is contentious. Here the effect of inflammation on activity of the NaHCO3 cotransporter, NBCe1, in the proximal colon was investigated. Interleukin10‐knockout (IL10‐/‐) mice infected with Helicobacter typhlonius, which develop colitis, were used as a model of inflammation, and uninfected, asymptomatic IL10‐/‐ mice served as controls. NBCe1 activity was measured as the 5‐(N‐ethyl‐N‐isopropyl) amiloride‐insensitive (EIPA, 20µM), 4,4'‐diisothiocyano‐2,2'‐stilbenedisulfonic acid‐sensitive (DIDS, 500µM), rate of intracellular pH recovery and acid extrusion (JH+) following acid loading of isolated crypts, while NBCe1 expression was measured by Western blotting and immunohistochemistry. In control tissues NBCe1 immunoreactivity was restricted to the surface cells and upper third of the crypts. Consistent with this there was little evidence of NBCe1 activity in the lower half of the crypts, with or without forskolin (10µM) stimulation. In contrast, in the surface cells, while NBCe1 was inactive under basal conditions (JH+EIPA=16.4±1.8mM.min‐1; JH+EIPA+DIDS=18.7±1.8 mM.min‐1), its activity was stimulated by forskolin (after forskolin JH+EIPA=24.5±2.7 mM.min‐1 and JH+EIPA+DIDS=13.6±1.1mM.min‐1). In inflamed tissues, total NBCe1 expression was reduced (P<0.001, n=6), there was no NBCe1 immunoreactivity in the surface cells and upper crypts, and forskolin did not stimulate NBCe1 activity in the surface cells. This suggests inflammation reduces colonic HCO3‐ secretion, which may affect luminal pH regulation and growth of luminal bacteria, possibly contributing to the dysbiosis seen in patients with inflammatory bowel disease, as well as modifying mucus hydration.Grant Funding Source: Supported by The Otago Medical Research Foundation
Bile acid chenodeoxycholate activation of Cl‐ transport involves epidermal growth factor receptor signal transduction in human colonic T84 cells (908.4)
Authors: Domingue Jada , Ao Mei , Sarathy Jayashree , Alrefai Waddah , Rao Mrinalini 
Published: June 2021

A role for EGFR signaling in BA action has been well described in colon cancer, but not as well in BA induced Cl‐ secretion. Our preliminary report (EB LB 10695, ‘13) showed that the EGFR inhibitor, AG1478, reduced CDCA (500 µM) mediated Cl‐ secretion by 69%, measured as short circuit current (Isc, µA.cm‐2), in T84 cells. Here we aim to further elucidate this mechanism. To determine if BA action involves matrix metalloproteinases (MMP), CDCA (500 µM) was added to T84 cells ± batimastat (BAT, 20 µM, 30’). CDCA itself increases Isc in 2‐3’ and the effect is sustained for > 20’. BAT inhibited the sustained phase (∆Isc: CDCA: 11.6±1.4; +BAT: 3.9±2.6, n=3). Interestingly the PKA inhibitor H89 has a similar effect (∆Isc: CDCA: 21±4.6; +H89 (30 µM): 1±0.3, n=5), suggesting that EGF and cAMP are involved in sustaining CDCA‐stimulated secretion. The distal steps of EGFR action do not involve Src or MEK as their respective inhibitors PP2 (10 µM, n=3), and PD98059 (20 µM, n=3) did not alter CDCA‐induced Isc. Immunoblotting showed CDCA (5’) increased EGFR (n=1), p38 (n=3), and ERK 1/2 (n=3) phosphorylation. Based on this data we postulate that CDCA stimulates Cl‐ secretion via MMP‐EGFR‐p38 signaling, but not via Src or the MEK‐ERK pathways. Together with our data (AJP 305:C447‐456, ‘13) that CDCA acts via PKA to activate CFTR, we summarize that CDCA evokes multiple signaling cascades and their cross talk needs further elucidation.Grant Funding Source: Supported by NIH and VA
The effects of insulin on gonadotropin‐releasing hormone secretion (911.1)
Authors: Herrera Danny , DiVall Sara , Wolfe Andrew 
Published: June 2021

The purpose of this study was to determine the role of insulin signaling in gonadotropin‐releasing hormone (GnRH) pulse generation. The Cre‐lox system was used to generate mice without insulin receptors in GnRH neurons (GnRH Cre+ IR fl/fl; GnRHIRKO). Hypothalami from female GnRHIRKO or littermate controls with intact insulin receptors in GnRH neurons (GnRH Cre‐), in the lean or diet induced obesity (DIO) state were incubated in static culture. Media collected every seven minutes was subjected to radioimmunoassay for GnRH. The pulse pattern in the control DIO state was visually less regular than other states. GnRH pulse amplitude was not different between GnRHIRKO or control mice in the lean or DIO state. Pulse frequency was trended lower in the control DIO state than in lean controls or GnRHIRKO lean or DIO hypothalami. However, this result was not statistically significant. Pulse interval was also longer in the control DIO state compared to the control lean or GnRHIRKO hypothalami. Insulin signaling in the GnRH neuron may play a role in GnRH pulse generation in the DIO but not the lean state.Grant Funding Source: Supported by: NIH K08HD056139 and APS Undergraduate Summer Research Fellowship
Endocrine disruption in endangered Oregon spotted frogs (Rana pretiosa) (911.9)
Authors: Griffing Aaron , Sessions Stanley 
Published: June 2021

The Oregon spotted frog (Rana pretiosa) is an endangered species residing in the Pacific Northwest whose dwindling numbers having caught the attention of biologists and environmental scientists. We tested the idea that R. pretiosa populations may be declining due to disruption of sex determination via endocrine disruption and/or parasite infection. Newly metamorphosed specimens were collected from localities in Oregon and their gonads were examined using paraffin histology. The sex ratio was 41% males and 59% females. This was not statistically significant. We found that 46% of the male frogs had oocytes developing in their testes. We also found that all specimens had numerous trematode cysts in their kidneys. These results from wild‐caught frogs were compared to an analysis of captive‐raised Wood Frogs (Rana sylvatica) from New York.Grant Funding Source: Supported by the Oregon Community Foundation and its grant
miRNAs as the molecular clock regulating mandibular osteoinduction (921.5)
Authors: Roberts Ryan , Merrill Amy 
Published: June 2021

Precise timing of cranial neural crest cell (CNCC) differentiation into bone is essential for proper formation of the craniofacial bones. Delayed osteoinduction leads to micrognathia, whereas premature osteoinduction causes craniosynostosis. To understand the etiology of these birth defects, we must uncover the molecular clock that regulates the onset of osteogenesis. CNCC autonomously control the timing of their differentiation into osteoblasts by regulating expression of Bone Morphogenetic Protein (BMP), yet how CNCC establish the timing of BMP expression remains unclear. Good candidates for intrinsic temporal control of gene expression are miRNAs, small non‐coding RNAs that target mRNAs for degradation. To identify specific miRNAs involved in mandibular osteogenesis, we performed microarray analysis on chick mandibles during the stages of osteoinduction. We identified and validated temporal expression changes in miRNAs whose targets include BMP signaling pathway members that are known to be involved in bone formation. For example, miR‐19b, which is upregulated at the start of osteogenesis, targets repressors of BMP signaling including Smurf1. Conversely, let‐7a and let‐7c, which are upregulated at the close of osteoinduction, target the BMP receptor AcvR‐2B. These results suggest that miRNAs intrinsically control the timing of CNCC osteoinduction by regulating the period of active BMP signaling.Grant Funding Source: NIH 5P30DE020750‐02
Molecular mechanisms that mediate mechanosensation during secondary chondrogenesis (921.9)
Authors: Woronowicz Katherine , Hague Molly , Schneider Richard 
Published: June 2021

The ability of cells to detect mechanical forces plays an essential role in generating the proper form and function of the craniofacial musculoskeletal system. One skeletal tissue that remains highly responsive to mechanical force is secondary cartilage, which develops along the surface of bone. Without appropriate mechanical stimulation, such as the forces that arise through motility of the embryo, secondary cartilage does not form, or degenerates. To elucidate how secondary cartilage is induced and maintained by mechanical force, we take advantage of the fact that secondary cartilage is closely associated with the specialized feeding of some species of birds. Duck utilize their jaws as a means to filter heavy sediment while chick peck at seed. Embryonic motility in duck induces secondary cartilage within the jaw adductor enthesis, creating a robust interface between tendon and bone on the coronoid process, and ultimately transducing the powerful muscle contractions needed to lift the jaw. In contrast, secondary cartilage is absent on the chick coronoid process. Our published and preliminary data lead us to hypothesize that species‐specific jaw anatomy generates dissimilar mechanical forces that differentially activate mechanosensitive Fibroblast Growth Factor and Calcium signaling pathways in duck versus chick, and instruct the duck jaw adductor enthesis to produce secondary cartilage. We employ gain‐ and loss‐of‐function approaches to modulate secondary chondrogenesis, as well as chick‐duck chimeras to test if chick cells can form secondary cartilage in a duck host environment. This research has the potential to translate into regenerative therapies to supplement surgical repair of torn and damaged muscle insertions.Grant Funding Source: Supported by NIH 5R01DE016402‐08
Expression and immunohistochemical localization of the Gβγ‐activated and calcineurin‐inhibited adenylyl cyclase isoforms in rat articular chondrocytes (922.9)
Authors: Memon Ismail , Khan Khalid , Siddiqui Sammer , Perveen Siddiqa , Ishaq Muhammad 
Published: June 2021

Aims: Cross‐talk between signal transduction pathways is essential to maintain normal homeostasis. cAMP mediated pathway is archetypical in mammalian cells and adenylyl cyclase (AC) is the effector enzyme that converts ATP into cAMP. We believe that AC/cAMP mediated pathway is involved in homeostasis of the articular cartilage. In this study we report the expression of Gβγ‐activated AC isoforms 2, 4, and 7 and calcineurin‐inhibited AC isoform 9 in rat articular chondrocytes.Main methods: Fresh slices of articular cartilage were taken from various synovial joints of rat and the expression of AC isoforms was determined by RT‐PCR. Immunohistochemistry was performed to localize these isoforms in articular chondrocytes.Key findings: All AC isoforms studied were found differentially expressed in different zones of the rat articular cartilage. Generally, expression of all AC isoforms increased with age.Significance: These data add to the information about signaling cascades possibly involved in articular chondrocytes. Presence of AC isoforms 2, 4, 7, and 9 suggest a role for the cAMP mediated 2nd messenger signaling cascades in articular chondrocytes. Further studies, such as measuring the AC and cAMP levels during development and differentiation of articular chondrocytes would confirm our results and help in understanding the signaling mechanisms in articular chondrocytes.Grant Funding Source: Supported by Seed money grant from the FHS‐Research Committee of the Aga Khan University, Karachi
A bologist's resource for plasmids: the DNASU and Protein Structure Initiative:Biology‐Materials repositories (928.1)
Authors: Seiler Catherine , Park Jin , Fiacco Michael , Throop Andrea , Steel Jason , Hunter Preston , LaBaer Joshua 
Published: June 2021

The Protein Structure Initiative:Biology‐Materials Repository (MR; http://psimr.asu.edu) sequence‐verifies, annotates, stores, and distributes plasmids created by the Protein Structure Initiative (PSI). The DNASU plasmid repository (http://dnasu.org), which houses the MR, distributes over 198,000 plasmids created by our laboratory, by various consortia and by individual laboratories for distributions. This collection includes expression vectors, plasmid containing over 14,000 human genes, and as of November 2013, over 81,000 protein expression plasmids from PSI:Biology Centers. Through our recently redesigned website, researchers can search or browse our collection using our five search options or six new browse features which include an interactive vector browse and a collection browse that displays plasmids annotated by SMART domains. Detailed information about each plasmid such as the full length sequence, vector information, a dynamic vector map, and associated publications can be found directly on the website. Each plasmid also links to external resources, including the PSI Structural Biology Knowledgebase (http://sbkb.org) facilitating cross‐referencing of plasmids to additional protein annotations and experimental data. All plasmids can be requested through DNASU, and our facility ships to researchers worldwide. With the added biological focus in PSI:Biology, the distribution of these materials will help researchers expand the knowledge of the role of proteins both in normal biological processes and in disease.Grant Funding Source: Supported by the National Institute of General Medical Sciences.
Elucidating the role of de novo mutations of cullin‐3 in hypertension (930.1)
Authors: Ibeawuchi Stella‐Rita , Agbor Larry , Quelle Frederick , Sigmund Curt 
Published: June 2021

Loss of Cullin3 (Cul3) ubiquitin ligase activity is linked to hypertension (HT) attributable to increased RhoA protein in vascular smooth muscle cells (VSMCs). Patients with de novo mutations that lead to genetic deletion of exon 9 in Cul3 (Cul3Δ9) have early onset HT, but the mechanistic basis for this effect is lacking. We hypothesize that the Cul3Δ9 mutation results in reduced RhoA ubiquitination and degradation, thus elevating activatable RhoA protein levels. In support of this hypothesis, we found that exogenous levels of RhoA were increased in HEK293T cells cotransfected with Cul3Δ9 and the adaptor protein, Bacurd1, compared to cells expressing full length Cul3 (Cul3WT). Both Cul3Δ9 and Cul3WT were neddylated and both bound to Bacurd1. However, Cul3Δ9 binding to E3 ubiquitin ligase, RBX1, was decreased compared to Cul3WT. Dimerization of Cul3 ubiquitin ligase complexes is required for efficient ubiquitination of some substrates, and we show that dimerization of Cul3Δ9 is significantly impaired compared to Cul3WT. Thus, we conclude that Cul3Δ9 retains some activities of Cul3WT, but defects in Cul3Δ9‐associated ubiquitin ligase activity may impair RhoA ubiquitination thus stabilizing RhoA protein. Future studies will investigate the in vivo consequence of Cul3Δ9 on blood pressure and vascular function in a transgenic mouse model expressing Cul3Δ9 specifically in the VSMCs.Grant Funding Source: Supported by NIH
Adaptor‐guided proteolysis initiates on the ribosome (932.1)
Authors: Puri Neha , Karzai Wali 
Published: June 2021

Adaptor proteins extend the substrate range and specificity of AAA+ proteases. The SspB adaptor protein enhances the efficiency of ClpXP mediated degradation of tmRNA‐tagged proteins. We have investigated the mechanism by which SspB captures tag‐bearing proteins. Our inquiries reveal that SspB associates with tmRNA‐rescued ribosomes in a trans‐translation dependent manner, in a process reliant on distinct recognition signals within the appended tmRNA degron. Specific alterations in the tmRNA peptide sequence or the peptide‐binding groove of SspB have adverse effects on the efficiency of SspB ribosome association and the subsequent degradation of marked proteins. Our studies have also unveiled the ability of SspB to capture additional nascent polypeptides that harbor genetically encoded yet degenerate forms of the tmRNA degron and modulate their intracellular levels through directed proteolysis. Based on these findings, we propose that SspB scans translating ribosomes to capture signal‐bearing proteins for guided proteolysis by the ClpXP protease.Grant Funding Source: Supported by NIH
Characterization of StiP, a starvation‐induced cysteine protease from Acinetobacter baylyi (933.4)
Authors: Daugherty Margaret , Lang Kristine , Lostroh C. Phoebe , Memmel Cody , Bernazzani Sarina , Reichert Blake , Dornbusch Amber , Arguella Joshua , Stanley Sarah 
Published: June 2021

We have investigated the gene ACIAD 1960 from Acinetobacter baylyi. A. baylyi is a soil bacterium that is of interest due to its ability to utilize a diverse set of nutrients as a source of carbon and nitrogen, and to its high level of competence with regards to natural transformation. Medically, it is becoming of interest as a potential source of hospital‐acquired infections. Previous work has shown that gene ACIAD1960 is expressed during long‐term stationary phase. We identified that the purified protein expressed by ACIAD 1960 encodes a cysteine protease activity. We named the cysteine protease “StiP” to reflect that it is a long‐term stationary phase induced protease. We present data from a deletion‐insertion mutation which demonstrates that lack of the protein causes alteration of normal cell morphology, smaller twitching zone size, and hypersensitivity to the oxyanion tellurite. The mutant may also have a small growth defect. We show that the presence of divalent metal ions differentially affect its activity. The presence of Ca2+, Mg2+ and Mn2+ all increase activity, with Mn2+ showing the greatest effect. The presence of Zn2+ significantly decreases activity. Additionally, we show that StiP activity decreases in the presence of reducing agents, suggesting that a disulfide bond maycontribute to its overall conformational stability and function.Grant Funding Source: Supported by Colorado College Natural Sciences Executive Committee grants to M.A.D., K.M.L., and C.
Reconstitution of the 26S proteasome reveals functional asymmetries in its AAA+ unfoldase (936.1)
Authors: Beckwith Robyn , Estrin Eric , Worden Evan , Martin Andreas 
Published: June 2021

The 26S proteasome is the major protease in eukaryotic cells responsible for selective protein degradation to mediate protein quality control and regulation, yet the detailed mechanisms by which the proteasomal heterohexameric AAA+ unfoldase drives ATP‐dependent protein degradation remain poorly understood. Delineating the roles of the six distinct ATPase subunits in substrate processing has been hindered by limitations in working with endogenous proteasomes due to misassembly or lethal degradation defects. We therefore developed a heterologous expression system to produce the unfoldase subcomplex from Saccharomyces cerevisiae in Escherichia coli and reconstituted the proteasome in vitro to perform systematic mutational analyses of the individual ATPase subunits. Our studies demonstrate that the six ATPases have distinct functions in degradation, corresponding to their positions in the spiral staircases adopted by their large AAA+ domains in the absence or presence of substrate. ATP hydrolysis in subunits at the top of the staircases is critical for substrate engagement and translocation. Whereas the unfoldase relies on this vertical asymmetry for substrate processing, interaction with the peptidase exhibits a pronounced three‐fold symmetry. Only three ATPase subunits, arranged in alternate positions within the unfoldase ring, contain a conserved C‐terminal hydrophobic/aromatic/unspecified (HbYX) motif that is critical for both peptidase binding and gate‐opening, whereas the C‐terminal tails of the interjacent ATPase subunits are dispensable. Our study provides an initial glimpse into the potential importance of the spiral staircase configurations of proteasomal ATPase subunits in substrate processing and highlights how the 26S proteasome may deviate from simpler, homomeric AAA+ proteases.Grant Funding Source: Supported by US National Institutes of Health grant R01‐GM094497‐01A1