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Cricetulus migratorius is widely distributed in the northwestern arid regions of China. Here, the complete mitochondrial genome (mitogenome) of C. migratorius is reported, to our knowledge, for the first time. It was found to be 16,246bp in length, including 13 protein-coding genes (PCGs), 2 rRNA genes, 22 tRNA genes, and one control region, and showed characteristics typical of the vertebrate mitogenome. Comparative analyses of mitogenomes of Cricetulus species showed that hamster mitogenomes had the same arrangement and organizational structure. The base composition was AT-rich for all Cricetulus species. We elaborated the PCG sequence and found that the main differences in the start and stop codons were in the ND3 gene; the most frequently used codons were AUU (Ile) and CUA (Leu), and the AT-content at the 3rd position was lower than that at 1st and 2nd positions in all Cricetulus species. The second tRNAser had lost the dihydrouridine stem-loop structure, which is found in all Cricetulus species. We also elucidated the structure of the control region (the extended termination associated sequences, the central conserved sequences, and the conserved sequence blocks), and the putative origin of replication for the light strand in all Cricetulus species. Based on mitogenome data with four reconstructed methods, phylogenetic trees showed high resolution in the divergent clades within Cricetulus. The results indicated that Cricetulus kamensis is at basal position, and has the earliest split among all Cricetulus species. Cricetulus griseus and Cricetulus longicaudatus clustered in a subclade, and were separate from C. kamensis. As previous studies have shown, Tscherskia triton is part of Cricetulus with high support value. It is noteworthy that C. migratorius is a young species, and has a close relationship with Allocricetulus eversmanni within Cricetulus, which is closer to the subgenus Allocricetulus.
Animals in the non-tropical zone usually demonstrate seasonal variations in immune function, which is important for their survival. In the present study, seasonal changes in immunity in striped hamsters (Cricetulus barabensis) were investigated to test the winter immunoenhancement hypothesis. Male hamsters were captured from the wild in the fall and winter of 2014 and in the spring and summer of 2015. Body mass, body fat mass and blood glucose levels of the hamsters were all highest in the summer, whereas relative fatness and thymus mass had no seasonal changes. Spleen mass was highest in the fall and white blood cells and phytohaemagglutinin (PHA) response indicative of cellular immunity were lowest in the summer among the four seasons, which supports the winter immunoenhancement hypothesis. IgG and IgM titers were lowest in the fall, which was against this hypothesis. Body fat mass had no correlations with cellular and humoral immunity, suggesting it was not the reason for seasonal changes in cellular and humoral immunity in males. Leptin titers were higher in spring and summer than in fall and winter. No correlation between leptin and cellular and humoral immunity suggested that leptin did not mediate their seasonal changes. Similarly, corticosterone levels were also higher in spring and summer than in fall and winter, which correlated negatively with cellular immunity but positively with IgG levels. This result implied that corticosterone has a suppressive effect on cellular immunity and an enhancing effect on humoral immunity. In summary, distinct components of immune systems exhibited different seasonal patterns. This article has an associated First Person interview with the first author of the paper.
The study was part of the UK EMS Genetic Toxicology trial, to determine the clastogenic potential of 4CMB in the bone-marrow cell chromosomes of the Chinese hamster. At dose levels of 50, 100, 200, 300 and 400 mg/kg of 4CMB statistical analysis of the data failed to show any significant differences in chromosomal damage between negative controls and 4CMB-treated groups. However, numerical increases in chromosomal damage were observed in the 4CMB-treated groups. The aberrations were at a low frequency but included the more severe types of chromosomal damage, such as exchange figures and dicentric chromosomes.
Photoperiod is an important factor of mammalian seasonal rhythm. The Harderian gland (HG) appears to act as a "standby" structure of the retinal-pineal axis, mediating light signals in vitro and neuroendocrine regulation in vivo; however, the effect of photoperiod on the HG is not clear. Here, we studied morphological differences in the HG of female striped dwarf hamsters (Cricetulus barabensis), a small mammal that experiences an annual rhythm, under different photoperiods (i.e., SP, short photoperiod; MP, moderate photoperiod; LP, long photoperiod), and further investigated the molecular mechanisms related to these morphological differences. Results showed that body weight, carcass weight, and HG weight were higher in the SP and LP groups than that in the MP group. Protein expression of hydroxyindole-o-methyltransferase, a key enzyme in melatonin synthesis, was higher in the SP group than in the other two groups. Somatostatin showed highest expression in the LP group. Furthermore, comparison of changes in the HG ultrastructure demonstrated autolysosome formation in the SP group. Protein aggregation and mRNA expression of LC3 and protein expression of LC3II/LC3I were higher in the SP group than in the MP group, indicating elevated autophagy under SP. Chromatin agglutination and mitochondrial damage were observed and bax/bcl2 and cytochrome C expression increased at the protein and mRNA levels in the SP and LP groups, suggesting increased apoptosis. Protein expression of dynamin-related protein 1 and mitochondrial fission factor (Mff) were highest in the SP group, suggesting elevated mitochondrial fission. Protein expression levels of adenosine triphosphate (ATP) synthase and citrate synthase were lower in the LP group than in the SP and MP groups. These results indicated that autophagy and apoptosis imbalance under SP and LP conditions may have led to HG weight loss and up-regulation of mitochondrial apoptosis may have weakened mitochondrial function under LP conditions. Finally, melatonin synthesis appeared to be positively correlated with the time hamsters entered darkness.
RFamide-related peptides (RFRPs) are orthologous to gonadotropin-inhibitory hormone (GnIH) inhibiting gonadotropin release. There are only two RFRP sequences (RFRP-1 and RFRP-3) encoded in rodents. RFRP-3, which was considered as a hypothetical inhibitor on GnRH, shows a stimulatory effect on the male Syrian and male Siberian hamster in short days. As a dominant rodent pest in northern China farmland, the striped hamster (Cricetulus barabensis) has higher reproductive activities and could act as a model to study the mechanism of reproduction. However, the effect of RFRP-3 on the reproductive activity for the striped hamster is less understood. In the study, we cloned 643 bp RFRP cDNA from the striped hamster hypothalamus, which contained an ORF of 570 bp encoding two RFamide-related peptide (RFRP) sequences: SPAPANKVPHSAANLPLRF-NH2 (C. barabensis RFRP-1) and TLSRVPSLPQRF-NH2 (C. barabensis RFRP-3). We also investigated the expression variation of RFRP mRNA and GnRH mRNA in the hypothalamus from hamsters with different developmental statuses (7-week-, 13-week- and 1.5-year-olds) using FQ-PCR, in which the 13-week-old female individuals were in estrous. The striped hamsters that are 7 weeks and 1.5 years old are non-breeding individuals, and those that are 13-week hamsters have breeding phenomena. The highest hypothalamus RFRP mRNA level was found in breeding males as compared to non-breeding males. Conversely, the lowest RFRP mRNA level in the hypothalamus was observed in breeding females, with no significant level when the breeding females were compared to the 7-week-old individuals. Additionally, the investigation of GnRH expression level showed a declining expression trend across the developmental stages (7-week-, 13-week- and 1.5-year-olds) in both sexes. Significant negative and positive relationships were detected in the 13-week estrous female (r=-0.997, P=0.035) and the 13-week male (r=0.998, P=0.029) striped hamsters respectively, which suggest that RFRP-3 has inhibitory and stimulatory effects on female and male adults respectively. Our results suggest that the effects of RFRP-3 on reproduction are sex- and developmental status-dependent in the striped hamster.
The SARS-CoV-2 pandemic has caused a yet unresolved global crisis. Effective medical intervention by vaccination or therapy seems to be the only possibility to control the pandemic. In this context, animal models are an indispensable tool for basic and applied research to combat SARS-CoV-2 infection. Here, we established a SARS-CoV-2 infection model in Chinese hamsters suitable for studying pathogenesis of the disease as well as pre-clinical testing of vaccines and therapies. This species of hamster is susceptible to SARS-CoV-2 infection as demonstrated by robust virus replication in the upper and lower respiratory tract accompanied by bronchitis and pneumonia as well as significant body weight loss following infection. The Chinese hamster features advantages compared to the Syrian hamster model, including more pronounced clinical symptoms, its small size, well-characterized genome, transcriptome and translatome data and availability of molecular tools.
The evolutionary process of an organism provides valuable data toward an understanding of the Earth evolution history. To investigate the relationship between the uplift of the Qinghai-Tibet Plateau (QTP) and mammalian evolution since the late Cenozoic, the geographic distribution of genetic variations in the Tibetan hamster Cricetulus kamensis was investigated using phylogeographical methods. In particular, population divergence, demographic history, genetic variation, and the prediction of species distribution area were investigated.
Both Cricetus cricetus and Phodopus sungorus mitochondrial genomes (mitogenomes) were sequenced and elaborated for the first time in the present study. Their mitogenomes contained 37 genes and showed typical characteristics of the vertebrate mitogenome. Comparative analysis of 10 cricetine mitogenomes indicated that they shared similar characteristics with those of other cricetines in terms of genes arrangement, nucleotide composition, codon usage, tRNA structure, nucleotide skew and the origin of replication of light strand. Phylogenetic relationship of the subfamily Cricetinae was reconstructed using mitogenomes data with the methods of Bayesian Inference and Maximum Likelihood. Phylogenetic analysis indicated that Cricetulus kamensis was at basal position and phylogenetically distant from all other Cricetulus species but had a close relationship with the group of Phodopus, and supported that the genus Urocricetus deserved as a separate genus rank. The phylogenetic status of Tscherskia triton represented a separate clade corresponding to a diversified cricetine lineage (Cricetulus, Allocricetulus, and Cricetus).
The integration of a range of technologies including microfluidics, surface-enhanced Raman scattering and confocal microspectroscopy has been successfully used to characterize in situ single living CHO (Chinese hamster ovary) cells with a high degree of spatial (in three dimensions) and temporal (1 s per spectrum) resolution. Following the introduction of a continuous flow of ionomycin, the real time spectral response from the cell was monitored during the agonist-evoked Ca(2+) flux process. The methodology described has the potential to be used for the study of the cellular dynamics of a range of signalling processes.
Unique anatomical and physiological features have made hamster species desirable research models. Comparative genomics and phylogenetic analysis of the hamster family members to clarify their evolution and genetic relationship, can provide a genetic basis for the comprehension of the variable research results obtained using different hamster models. The Syrian golden hamster (Mesocricetus auratus) is the most widely used species. In this study, we sequenced the complete mitochondrial genome (mitogenome) of M. auratus, compared it with the mitogenome of other Cricetinae subfamily species, and defined its phylogenetic position in the Cricetidae family. Our results show that the mitogenome organization, gene arrangement, base composition, and genetic analysis of the protein coding genes (PCGs) of M. auratus are similar to those observed in previous reports on Cricetinae species. Nonetheless, our analysis clarifies some striking differences of M. auratus relative to other subfamily members, namely distinct codon usage frequency of TAT (Tyr), AAT (Asn), and GAA (Glu) and the presence of the conserved sequence block 3 (CSB-3) in the control region of M. auratus mitogenome and other hamsters (not found in Arvicolinae). These results suggest the particularity of amino acid codon usage bias of M. auratus and special regulatory signals for the heavy strand replication in Cricetinae. Additionally, Bayesian inference/maximum likelihood (BI/ML) tree shows that Cricetinae and Arvicolinae are sister taxa sharing a common ancestor, and Neotominae split prior to the split between Cricetinae and Arvicolinae. Our results support taxonomy revisions in Cricetulus kamensis and Cricetulus migratorius, and further revision is needed within the other two subfamilies. Among the hamster research models, Cricetulus griseus is the species with highest sequence similarity and closer genetic relationship with M. auratus. Our results show mitochondrial DNA diversity of M. auratus and other Cricetinae species and provide genetic basis for judgement of different hamster models, promoting the development and usage of hamsters with regional characteristics.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the recent COVID-19 public health crisis. Bat is the widely believed original host of SARS-CoV-2. However, its intermediate host before transmitting to humans is not clear. Some studies proposed pangolin, snake, or turtle as the intermediate hosts. Angiotensin-converting enzyme 2 (ACE2) is the receptor for SARS-CoV-2, which determines the potential host range for SARS-CoV-2. On the basis of structural information of the complex of human ACE2 and SARS-CoV-2 receptor-binding domain (RBD), we analyzed the affinity to S protein of the 20 key residues in ACE2 from mammal, bird, turtle, and snake. Several ACE2 proteins from Primates, Bovidae, Cricetidae, and Cetacea maintained the majority of key residues in ACE2 for associating with SARS-CoV-2 RBD. The simulated structures indicated that ACE2 proteins from Bovidae and Cricetidae were able to associate with SARS-CoV-2 RBD. We found that nearly half of the key residues in turtle, snake, and bird were changed. The simulated structures showed several key contacts with SARS-CoV-2 RBD in turtle and snake ACE2 were abolished. This study demonstrated that neither snake nor turtle was the intermediate hosts for SARS-CoV-2, which further reinforced the concept that the reptiles are resistant against infection of coronavirus. This study suggested that Bovidae and Cricetidae should be included in the screening of intermediate hosts for SARS-CoV-2.
The metazoan endoplasmic reticulum (ER) serves both as a hub for maturation of secreted proteins and as an intracellular calcium storage compartment, facilitating calcium-release-dependent cellular processes. ER calcium depletion robustly activates the unfolded protein response (UPR). However, it is unclear how fluctuations in ER calcium impact organellar proteostasis. Here, we report that calcium selectively affects the dynamics of the abundant metazoan ER Hsp70 chaperone BiP, by enhancing its affinity for ADP. In the calcium-replete ER, ADP rebinding to post-ATP hydrolysis BiP-substrate complexes competes with ATP binding during both spontaneous and co-chaperone-assisted nucleotide exchange, favouring substrate retention. Conversely, in the calcium-depleted ER, relative acceleration of ADP-to-ATP exchange favours substrate release. These findings explain the rapid dissociation of certain substrates from BiP observed in the calcium-depleted ER and suggest a mechanism for tuning ER quality control and coupling UPR activity to signals that mobilise ER calcium in secretory cells.
The cotton rat (Sigmodon hispidus) belongs to the rodent family of Cricetidae and provides a powerful model to study the pathogenesis of human respiratory viruses and measles virus. Recent studies in other rodent models have suggested a role for invariant natural killer T (iNKT) cells in antiviral immunity and vaccination against respiratory virus infections. Using new experimental tools, we provide the first evidence for a functional CD1d cell molecule (crCD1d) and iNKT T-cell receptor in cotton rats. The crCD1d cDNA sequence was identified and crCD1d transductants showed that monoclonal antibody WTH-2 stains crCD1d as efficiently as mouse or rat CD1d. The expression of crCD1d was clearly weaker for thymocytes and B cells, and higher for T cells, which is different to what is found in murine species. The antigen-presenting capacity of crCD1d was demonstrated with crCD1d-immunoglobulin dimers loaded with the glycolipid PBS57, which bound iNKT T-cell receptors. Evidence for functional cotton rat iNKT cells was provided by detection of interferon-γ and interleukin-4 in cultures of splenocytes stimulated with PBS57 and α-galactosylceramide and by specific staining of about 0·2% of splenocytes with PBS57-loaded crCD1d dimers. Canonical AV14/AJ18 rearrangements were identified and found to contain multiple members of the AV14 (AV11) family. One of them was expressed and found to bind CD1d dimers. In summary, these data provide the first evidence for functional CD1d molecules and iNKT T-cell receptors in cotton rats and provide the tools to analyse them both in the cotton rat model of infectious diseases.
A novel I2/TBHP mediated domino synthesis of 2-(2,4-dioxo-1,4-dihydroquinazolin-3(2H)-yl)-N-phenyl benzamides by reaction of isatins with o-amino N-aryl/alkyl benzamides was described. This was the first application of o-amino N-aryl/alkyl benzamides participating in oxidative rearrangement with isatins for synthesis of desired products. The synthesized compounds contained amide and quinazoline units and their combination resulted in molecular hybridization of two important pharmacophores. In this study, the synthesized compounds 3a-r were screened for cytotoxicity against four cancer cell lines A549, DU145, B16-F10, and HepG2 and also non-cancerous cell line CHO-K1. The compounds 3c, 3l and 3o gave promising results. The in silico molecular docking studies (PDB ID 1N37) also validated the anticancer activity of these compounds showing good binding affinity with target DNA and by acting as DNA intercalators.
Monoclonal antibody (mAb) interchain disulfide bond reduction can cause a loss of function and negatively impact the therapeutic's efficacy and safety. Disulfide bond reduction has been observed at various stages during the manufacturing process, including processing of the harvested material. The factors and mechanisms driving this phenomenon are not fully understood. In this study, we examined the host cell proteome as a potential factor affecting the susceptibility of a mAb to disulfide bond reduction in the harvested cell culture fluid (HCCF). We used untargeted liquid-chromatography-mass spectrometry-based proteomics experiments in conjunction with a semi-automated protein identification workflow to systematically compare Chinese hamster ovary (CHO) cell protein abundances between bioreactor conditions that result in reduction-susceptible and reduction-free HCCF. Although the growth profiles and antibody titers of these two bioreactor conditions were indistinguishable, we observed broad differences in host cell protein (HCP) expression. We found significant differences in the abundance of glycolytic enzymes, key protein reductases, and antioxidant defense enzymes. Multivariate analysis of the proteomics data determined that upregulation of stress-inducible endoplasmic reticulum (ER) and other chaperone proteins is a discriminatory characteristic of reduction-susceptible HCP profiles. Overall, these results suggest that stress response pathways activated during bioreactor culture increase the reduction-susceptibility of HCCF. Consequently, these pathways could be valuable targets for optimizing culture conditions to improve protein quality.
Unlike most kinesins, mitotic centromere-associated kinesin (MCAK) does not translocate along the surface of microtubules (MTs), but instead depolymerizes them. Among the motile kinesins, refinements that are unique for specific cellular functions, such as directionality and processivity, are under the control of a "neck" domain adjacent to the ATP-hydrolyzing motor domain. Despite its apparent lack of motility, MCAK also contains a neck domain. We found that deletions and alanine substitutions of highly conserved positively charged residues in the MCAK neck domain significantly reduced MT depolymerization activity. Furthermore, substitution of MCAK's neck domain with either the positively charged KIF1A K-loop or poly-lysine rescues the loss of MT-depolymerizing activity observed in the neckless MCAK mutant. We propose that the neck, analogously to the K-loop, interacts electrostatically with the tubulin COOH terminus to permit diffusional translocation of MCAK along the surface of MTs. This weak-binding interaction may also play an important role in processivity of MCAK-induced MT depolymerization.
Copaifera species folkloric names are "copaíbas, copaibeiras, copaívas or oil stick", which are widely used in Brazilian folk medicine. Among all ethnopharmacological applications described for Copaifera spp oleoresins, their anti-inflammatory effect stands out. However, the knowledge of anti-inflammatory and antinociceptive properties of Copaifera pubiflora Benth is scarce.
The production of recombinant proteins in mammalian cell lines is one of the most important areas in biopharmaceutical industry. Viral transcriptional promoters are widely used to express recombinant proteins in mammalian cell lines. However, these promoters are susceptible to silencing, thus limiting protein productivity. Some CpG islands can avoid the silencing of housekeeping genes; for that reason, they have been used to increase the production of recombinant genes in cells of animal origin. In this study, we evaluated the CpG island of the promoter region of the β-actin gene of Cricetulus griseous (Chinese hamster), associated to the Cytomegalovirus (CMV) promoter, to increase recombinant antibodies production in Chinese Hamster Ovary (CHO) cells.
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