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Novel insights into autophagosome biogenesis revealed by cryo‐electron tomography
Recent studies utilising cryo-electron tomography and detailed analysis of the image data have revealed novel information on the membrane dynamics of autophagosome biogenesis, including the shape and dimensions of omegasomes, phagophores and autophagosomes, and their relationships with the organelles around them. This review summarises the findings of three recent papers revealing new exciting information on phagophore biogenesis.
Autophagosome biogenesis, from the appearance of the phagophore to elongation and closure into an autophagosome, is one of the long-lasting open questions in the autophagy field. Recent studies utilising cryo-electron tomography and detailed analysis of the image data have revealed new information on the membrane dynamics of these events, including the shape and dimensions of omegasomes, phagophores and autophagosomes, and their relationships with the organelles around them. One of the important predictions from the new results is that 60–80% of the autophagosome membrane area is delivered by direct lipid transfer or lipid synthesis. Cryo-electron tomography can be expected to provide new directions for autophagy research in the near future.
Modulatory effects of point‐mutated IL‐32θ (A94V) on tumor progression in triple‐negative breast cancer cells
IL-32θ (A94V) inhibits phosphorylation of FAK and IκBα. IL-32θ (A94V) inhibits the expression and translocation of β-catenin by inhibiting phosphorylated FAK. Additionally, NF-κB is inhibited by IL-32θ (A94V) via the suppression of phosphorylated IκBα. Thus, IL-32θ (A94V) reduces migration, proliferation, and inflammation in breast cancer via the FAK-PI3K-GSK3 and NF-κB pathways.
Abstract
Breast cancer is a frequently diagnosed cancer and the leading cause of death among women worldwide. Tumor-associated macrophages stimulate cytokines and chemokines, which induce angiogenesis, metastasis, proliferation, and tumor-infiltrating immune cells. Although interleukin-32 (IL-32) has been implicated in the development and modulation of several cancers, its function in breast cancer remains elusive. Mutation of interleukin-32θ (IL-32θ) in the tissues of patients with breast cancer was detected by Sanger sequencing. RT-qPCR was used to detect the mRNA levels of inflammatory cytokines, chemokines, and mediators. The secreted proteins were detected using respective enzyme-linked immunosorbent assays. Evaluation of the inhibitory effect of mutant IL-32θ on proliferation, migration, epithelial–mesenchymal transition (EMT), and cell cycle arrest in breast cancer cells was conducted using MTS assays, migration assays, and Western blotting. A point mutation (281C>T, Ala94Val) was detected in IL-32θ in both breast tumors and adjacent normal tissues, which suppressed the expression of pro-inflammatory factors, EMT factors, and cell cycle related factors. Mutated IL-32θ inhibited the expression of inflammatory factors by regulating the NF-κB pathway. Furthermore, mutated IL-32θ suppressed EMT markers and cell cycle related factors through the FAK/PI3K/AKT pathway. It was inferred that mutated IL-32θ modulates breast cancer progression. Mutated IL-32θ (A94V) inhibited inflammation, EMT, and proliferation in breast cancer by regulating the NF-κB (p65/p50) and FAK-PI3K-GSK3 pathways.
A novel perspective on the role of nucleus accumbens neurons in encoding associative learning
The nucleus accumbens is key for encoding reward/aversion and associative learning, being the limbic-motor interface of the brain. This encoding occurs through activity of medium spiny neurons (MSNs) that express either dopamine receptor D1 or D2. Here, we discuss evidence supporting a complex and complementary role of D1- and D2-MSNs in encoding both appetitive and aversive cue–outcome associative learning.
The nucleus accumbens (NAc) has been considered a key brain region for encoding reward/aversion and cue–outcome associations. These processes are encoded by medium spiny neurons that express either dopamine receptor D1 (D1-MSNs) or D2 (D2-MSNs). Despite the well-established role of NAc neurons in encoding reward/aversion, the underlying processing by D1-/D2-MSNs remains largely unknown. Recent electrophysiological, optogenetic and calcium imaging studies provided insight on the complex role of D1- and D2-MSNs in these behaviours and helped to clarify their involvement in associative learning. Here, we critically discuss findings supporting an intricate and complementary role of NAc D1- and D2-MSNs in associative learning, emphasizing the need for additional studies in order to fully understand the role of these neurons in behaviour.
Biochemical and cellular studies of three human 3‐phosphoglycerate dehydrogenase variants responsible for pathological reduced L‐serine levels
In the brain, L-serine is produced through the phosphorylated pathway (PP). hPHGDH catalyzes the first and rate-limiting step in the PP. Three variants related to hPHGDH deficiency and Neu-Laxova syndrome have been studied. V261M, V425M, and V490M substitutions alter the kinetic and structural properties of hPHGDH. Variants ectopic expression results in protein aggregation and reduced L-serine level.
Abstract
In the brain, the non-essential amino acid L-serine is produced through the phosphorylated pathway (PP) starting from the glycolytic intermediate 3-phosphoglycerate: among the different roles played by this amino acid, it can be converted into D-serine and glycine, the two main co-agonists of NMDA receptors. In humans, the enzymes of the PP, namely phosphoglycerate dehydrogenase (hPHGDH, which catalyzes the first and rate-limiting step of this pathway), 3-phosphoserine aminotransferase, and 3-phosphoserine phosphatase are likely organized in the cytosol as a metabolic assembly (a “serinosome”). The hPHGDH deficiency is a pathological condition biochemically characterized by reduced levels of L-serine in plasma and cerebrospinal fluid and clinically identified by severe neurological impairment. Here, three single-point variants responsible for hPHGDH deficiency and Neu-Laxova syndrome have been studied. Their biochemical characterization shows that V261M, V425M, and V490M substitutions alter either the kinetic (both maximal activity and K m for 3-phosphoglycerate in the physiological direction) and the structural properties (secondary, tertiary, and quaternary structure, favoring aggregation) of hPHGDH. All the three variants have been successfully ectopically expressed in U251 cells, thus the pathological effect is not due to hindered expression level. At the cellular level, mistargeting and aggregation phenomena have been observed in cells transiently expressing the pathological protein variants, as well as a reduced L-serine cellular level. Previous studies demonstrated that the pharmacological supplementation of L-serine in hPHGDH deficiencies could ameliorate some of the related symptoms: our results now suggest the use of additional and alternative therapeutic approaches.
Genetic lineage tracing in skin reveals predominant expression of HEY2 in dermal papilla during telogen and that HEY2+ cells contribute to the regeneration of dermal cells during wound healing
Induction of reporter expression in Hey2-CreERT2, Rosa26-ZsGreen mice reveals that Hey2 is predominantly expressed in dermal papilla (DP) cells in skin. Lineage tracing of these HEY2+ cells during wound healing indicates that HEY2+ cells exhibit stemness properties and can contribute to the regeneration of dermal cells.
Dermal papilla (DP) cells are specialized mesenchymal cells that play a crucial role in regulating hair morphology, colour and growth through the secretion of specific factors. It is still unclear what the source of progenitor cells is for dermal cell regeneration during wound healing, and whether DP cells are involved in this process. We analyzed the gene expression profile of various skin cell populations using existing datasets and found that the Hey2 gene was predominantly expressed in DP cells. We introduced Hey2-CreERT2 knockin mice and crossed them with Rosa26-ZsGreen reporter mice. After induction in the double transgenic mice by administration of tamoxifen, the reporter ZsGreen was found to be predominantly expressed in DP cells both at anagen and telogen phases, and broadly expressed in some other dermal cells at anagen. We also created a wound after tamoxifen induction, and found there were abundant ZsGreen+ cells in the regenerated dermis. We conclude that the HEY2+ DP cells and dermal cells exhibit some stemness properties and can contribute to the dermal cell regeneration during wound healing.
Downregulation of microRNA‐326 enhances ZNF322A expression, transcriptional activity and tumorigenic effects in lung cancer
Oncogenic ZNF322A transcription factor is overexpressed in lung cancer. Downregulated miR-326 promotes ZNF322A-induced tumor growth and metastasis. This study reveals that miR-326-low/ZN322A-high profile is a biomarker to predict poor prognosis in lung cancer.
Abstract
Zinc finger protein ZNF322A is an oncogenic transcription factor. Overexpression of ZNF322A activates pro-metastasis, cancer stemness, and neo-angiogenesis-related genes to enhance lung cancer progression. However, the upstream regulator of ZNF322A is not well defined. Dysregulation of microRNAs (miRNAs) can mediate cancer cell growth, migration, and invasion to promote tumorigenesis. Here, we uncover the mechanism of miRNA-mediated transcriptional regulation in ZNF322A-driven oncogenic events. ZNF322A harbors several putative miRNA-binding sites in the 3′-untranslated region (UTR). We validated that miR-326 downregulated ZNF322A-3′-UTR luciferase activity and mRNA expression. Furthermore, miR-326 suppressed the expression of ZNF322A-driven cancer-associated genes such as cyclin D1 and alpha-adducin. Reconstitution experiments by ectopic overexpression of ZNF322A abolished miR-326-suppressed cancer cell proliferation and cell migration capacity. Moreover, miR-326 attenuated ZNF322A-induced tumor growth and lung tumor metastasis in vivo. Clinically, the expression of miR-326 negatively correlated with ZNF322A mRNA expression in surgically resected tissues from 120 non-small cell lung cancer (NSCLC) patients. Multivariate Cox regression analysis demonstrated that NSCLC patients with low miR-326/high ZNF322A profile showed poor overall survival. Our results reveal that the deregulated expression of miR-326 leads to hyperactivation of ZNF322A-driven oncogenic signaling. Targeting the miR-326/ZNF322A axis would provide new therapeutic strategies for lung cancer patients.
Coordinating nucleoporin condensation and nuclear pore complex assembly
The nuclear pore complex is a large multiprotein complex traversing the nuclear envelope. Many of its components harbor intrinsically disordered regions that undergo spontaneous condensation. Here, we discuss how assembly factors may guide their condensation, which must be carefully coordinated in space and time. We further discuss how defects in this process contribute to human pathologies, focusing on neurological disorders.
The nuclear pore complex (NPC) is among the most elaborate protein complexes in eukaryotes. While ribosomes and proteasomes are known to require dedicated assembly machinery, our understanding of NPC assembly is at a relatively early stage. Defects in NPC assembly or homeostasis are tied to movement disorders, including dystonia and amyotrophic lateral sclerosis (ALS), as well as aging, requiring a better understanding of these processes to enable therapeutic intervention. Here, we discuss recent progress in the understanding of NPC assembly and highlight how related defects in human disorders can shed light on NPC biogenesis. We propose that the condensation of phenylalanine-glycine repeat nucleoporins needs to be carefully controlled during NPC assembly to prevent aberrant condensation, aggregation, or amyloid formation.
Advances in screening, synthesis, modification, and biomedical applications of peptides and peptide aptamers
This comprehensive review explores the latest advancements in the screening, synthesis, modification, and biomedical applications of peptides and peptide aptamers. It discusses screening techniques, diverse synthesis strategies, and various modification approaches employed to enhance their properties. The review also highlights the broad range of biomedical applications where peptides and peptide aptamers have shown promise, including drug delivery, therapeutics, diagnostics, and biomaterials.
Abstract
Peptides and peptide aptamers have emerged as promising molecules for a wide range of biomedical applications due to their unique properties and versatile functionalities. The screening strategies for identifying peptides and peptide aptamers with desired properties are discussed, including high-throughput screening, display screening technology, and in silico design approaches. The synthesis methods for the efficient production of peptides and peptide aptamers, such as solid-phase peptide synthesis and biosynthesis technology, are described, along with their advantages and limitations. Moreover, various modification techniques are explored to enhance the stability, specificity, and pharmacokinetic properties of peptides and peptide aptamers. This includes chemical modifications, enzymatic modifications, biomodifications, genetic engineering modifications, and physical modifications. Furthermore, the review highlights the diverse biomedical applications of peptides and peptide aptamers, including targeted drug delivery, diagnostics, and therapeutic. This review provides valuable insights into the advancements in screening, synthesis, modification, and biomedical applications of peptides and peptide aptamers. A comprehensive understanding of these aspects will aid researchers in the development of novel peptide-based therapeutics and diagnostic tools for various biomedical challenges.
Preece–Baines 1 model validation for cross‐sectional data in male soccer players according to maturity status
Abstract
Aim
The aim of the present study was to compare the Preece–Baines 1 (PB1) model fit between longitudinal and cross-sectional data in male soccer players and to adjust the height growth curve by maturity status.
Methods
A final sample of 57 male Portuguese soccer players from professional soccer academies was included. Longitudinal height records were measured between 8 and 17 times in each subject from 2–8 years to 14–17 years. Additionally, longitudinal height records were used as cross-sectional data along with 1087 cross-sectional height records taken from 602 Portuguese soccer players. Skeletal age was estimated by Tanner-Whitehouse III method from a left hand–wrist radiograph. Age at peak height velocity (PHV) was estimated by PB1 model for longitudinal and cross-sectional data and by maturity status.
Results
No significant differences were found between all the longitudinal estimates of 57 players and the random cross-sectional samples for, S1 parameter and for growth velocity at PHV, at TO, and for age at PHV. The age at PHV in early, on-time, and late maturers were 12.26, 12.9, and 13.58 years, respectively.
Conclusion
PB1 adjusted the height growth of Portuguese male soccer players from cross-sectional data, obtaining an estimate PHV very similar to that found from longitudinal data. A maturity time difference of ≈0.6 years was found between the age at PHV of on-time, early, and on-time and late maturity state.