Non-coding RNA regulates phage sensitivity in Listeria monocytogenes

Non-coding RNAs (ncRNAs) have gained increasing attention as their diverse roles in virulence and environmental stress in Listeria monocytogenes have become clearer. The ncRNA rliB is an atypical member of the CRISPR family, conserved at the same genomic locus in all analyzed L. monocytogenes genomes and also in other Listeria species. In this study, rliB defective mutants (Lm3-22-ΔrliB) were constructed by homologous recombination. The growth cycle of Lm3-22-ΔrliB mutants was slower than that of wild-type Lm3-22. The sensitivity of Lm3-22-ΔrliB to the Listeria phage vB-LmoM-SH3-3 was significantly increased, and the efficiency of plaque formation was enhanced by 128 fold. Compared with wild type, the adhesion and invasion of Lm3-22-ΔrliB decreased significantly (9.3% and 1.33%, respectively). After 4 hours of infection, the proliferation of Lm3-22-ΔrliB in RAW264.7 cells also decreased significantly. Transcription level of invasion-related surface proteins showed that the internalin genes lmo0610 and lm0514, and the peptidoglycan binding protein gene lmo1799 in Lm3-22-ΔrliB were significantly increased. In addition, after interaction with phage, the transcription levels of inlA, lmo0610, lmo1799, lmo2085, and lmo0514 in Lm3-22-ΔrliB cells were significantly upregulated, while inlB was downregulated, compared with Lm3-22 control group with phage treatment. Therefore, rliB deletion effectively regulated the interaction between Listeria and phage, weaken its invasion ability, and provided a new theoretical basis for biocontrol of phage.

Synaptotagmin-13 Is a Neuroendocrine Marker in Brain, Intestine and Pancreas

Synaptotagmin-13 (Syt13) is an atypical member of the vesicle trafficking synaptotagmin protein family. The expression pattern and the biological function of this Ca2+-independent protein are not well resolved. Here, we have generated a novel Syt13-Venus fusion (Syt13-VF) fluorescence reporter allele to track and isolate tissues and cells expressing Syt13 protein. The reporter allele is regulated by endogenous cis-regulatory elements of Syt13 and the fusion protein follows an identical expression pattern of the endogenous Syt13 protein. The homozygous reporter mice are viable and fertile. We identify the expression of the Syt13-VF reporter in different regions of the brain with high expression in tyrosine hydroxylase (TH)-expressing and oxytocin-producing neuroendocrine cells. Moreover, Syt13-VF is highly restricted to all enteroendocrine cells in the adult intestine that can be traced in live imaging. Finally, Syt13-VF protein is expressed in the pancreatic endocrine lineage, allowing their specific isolation by flow sorting. These findings demonstrate high expression levels of Syt13 in the endocrine lineages in three major organs harboring these secretory cells. Collectively, the Syt13-VF reporter mouse line provides a unique and reliable tool to dissect the spatio-temporal expression pattern of Syt13 and enables isolation of Syt13-expressing cells that will aid in deciphering the molecular functions of this protein in the neuroendocrine system.

An atypical NLR protein modulates the NRC immune receptor network

The NRC immune receptor network has evolved in asterid plants from a pair of linked genes into a genetically dispersed and phylogenetically structured network of sensor and helper NLR (nucleotide-binding domain and leucine-rich repeat-containing) proteins. In some species, such as the model plant Nicotiana benthamiana and other Solanaceae, the NRC network forms up to half of the NLRome, and NRCs are scattered throughout the genome in gene clusters of varying complexities. Here, we describe NRCX, an atypical, but essential member of the NRC family that lacks canonical features of these NLR helper proteins, such as a functional N-terminal MADA motif and the capacity to trigger autoimmunity. In contrast to other NRCs, systemic gene silencing of NRCX markedly impairs plant growth resulting in a dwarf phenotype. Remarkably, dwarfism of NRCX silenced plants is partially dependent on NRCX paralogs NRC2 and NRC3, but not NRC4. Despite its negative impact on plant growth when silenced systemically, transient RNA interference of NRCX in mature N. benthamiana leaves does not result in visible cell death phenotypes. However, alteration of NRCX expression modulates the hypersensitive response mediated by NRC2 and NRC3 in a manner consistent with a negative role for NRCX in the NRC network. We conclude that NRCX is an atypical member of the NRC network that has evolved to contribute to the homeostasis of this genetically unlinked NLR network.

Post-translational modifications of CDK5 and their biological roles in cancer

Post-translational modifications (PTMs) of Cyclin-dependent kinase 5 (CDK5) have emerged as important regulatory mechanisms that modulate cancer development in patients. Though CDK5 is an atypical member of the cyclin-dependent kinase family, its aberrant expression links to cell proliferation, DNA damage response, apoptosis, migration and angiogenesis in cancer. Current studies suggested that, new PTMs on CDK5, including S-nitrosylation, sumoylation, and acetylation, serve as molecular switches to control the kinase activity of CDK5 in the cell. However, a majority of these modifications and their biological significance in cancer remain uncharacterized. In this review, we discussed the role of PTMs on CDK5-mediated signaling cascade, and their possible mechanisms of action in malignant tumors, as well as the challenges and future perspectives in this field. On the basis of the newly identified regulatory signaling pathways of CDK5 related to PTMs, researchers have investigated the cancer therapeutic potential of chemical compounds, small-molecule inhibitors, and competitive peptides by targeting CDK5 and its PTMs. Results of these preclinical studies demonstrated that targeting PTMs of CDK5 yields promising antitumor effects and that clinical translation of these therapeutic strategies is warranted.

Synaptotagmin13 Is A Neuroendocrine Marker In Brain, Intestine And Pancreas

Synaptotagmin13 (Syt13) is an atypical member of the vesicle trafficking Synaptotagmin protein family. The expression pattern and the biological function of this Ca2+-independent protein are not well resolved. Here, we have generated a novel Syt13-Venus fusion (Syt13-VF) fluorescence reporter allele to track and isolate tissues and cells expressing Syt13 protein. The reporter allele is regulated by endogenous cis-regulatory elements of Syt13 and the fusion protein follows an identical expression pattern of the endogenous Syt13 protein. The homozygous reporter mice are viable and fertile. We identify the expression of the Syt13-VF reporter in different regions of the brain with high expression in tyrosine hydroxylase (TH)-expressing and oxytocin-producing neuroendocrine cells. Moreover, Syt13-VF is highly restricted to all enteroendocrine cells in the adult intestine that can be traced in live imaging. Finally, Syt13-VF protein is expressed in the pancreatic endocrine lineage allowing their specific isolation by flow sorting. These findings demonstrate high expression levels of Syt13 in the endocrine lineages in three major organs harboring these secretory cells. Collectively, the Syt13-VF reporter mouse line provides a unique and reliable tool to dissect the spatio-temporal expression pattern of Syt13 and enables isolation of Syt13-expressing cells that will aid in deciphering the molecular functions of this protein in the neuroendocrine system.

The Rise and Fall of Bishop Georg Wolfgang Chiolich, 1699–1764

In my prosopographic examination of new aristocrats in the eighteenth century, I came across Georg Wolfgang Chiolich, an atypical member of this group, as he was the only one to receive the title of baron for himself alone, as a bishop, during the century. What was the point of conferring a noble title on a bishop? Searching for a reply to this question, we can gain some insight into the social, economic, and especially political processes of the Habsburg Empire in the mid-eighteenth century. Descended from a wealthy patrician family of Senj (Zengg), while Chiolich proved to be a talented organizer and took significant steps towards rebuilding his still-ruined bishopric in the mid-eighteenth century, he may not have been a saint. The rumors about the bishop of Senj (Zengg) finally escalated into a scandal in Vienna and in the Holy See in 1759, when a local noble family accused him of making a daughter of the head of the family pregnant. The legal proceedings, including the investigative material, were partly preserved in the Vatican Archives, and most of them were published by Tihamér Vanyó. Georg Wolfgang Chiolich eventually traveled to Rome, where the investigation declared him innocent despite all the efforts of the affronted family. If we look more closely at the main stages of his career and the course of this investigation, I believe that we can get closer to the political, social and economic conditions of a peripheral region of the Kingdom of Hungary.

The Role of RhoH in TCR Signalling and Its Involvement in Diseases

As an atypical member of the Rho family small GTPases, RhoH shares less than 50% sequence similarity with other members, and its expression is commonly observed in the haematopoietic lineage. To date, RhoH function was observed in regulating T cell receptor signalling, and less is known in other haematopoietic cells. Its activation may not rely on the standard GDP/GTP cycling of small G proteins and is thought to be constitutively active because critical amino acids involved in GTP hydrolysis are absent. Alternatively, its activation can be regulated by other types of regulation, including lysosomal degradation, somatic mutation and transcriptional repressor, which also results in an altered protein expression. Aberrant protein expression of RhoH has been implicated not only in B cell malignancies but also in immune-related diseases, such as primary immunodeficiencies, systemic lupus erythematosus and psoriasis, wherein its involvement may provide the link between immune-related diseases and cancer. RhoH association with these diseases involves several other players, including its interacting partner, ZAP−70; activation regulators, Vav1 and RhoGDI and other small GTPases, such as RhoA, Rac1 and Cdc42. As such, RhoH and its associated proteins are potential attack points, especially in the treatment of cancer and immune-related diseases.

Investigating the determinants of mammalian Mastl kinase activation

ABSTRACTMastl (Greatwall) kinase is an essential mitotic protein kinase. Mastl is an atypical member of AGC family with a unique long stretch of non-conserved middle region. The mechanism of its phosphorylation dependent activation has been studied in Xenopus egg extracts, revealing several phosphosites that were suggested to be crucial for kinase activation. These residues correspond to T193 and T206 in the activation loop, and S861 in the C-tail of mouse Mastl. By combining a chemically inducible knockout system to deplete the endogenous Mastl and a viral expression system to ectopically express the mutant variants, we obtained a viable knockout clone that expresses the S861A and S861D mutants. We observed that proliferation rates of the MastlS861A and MastlS861D clones were comparable. Our results have revealed that phosphorylation of the turn motif phosphosite (S861) is auxiliary and it is not indispensable for Mastl function.