The Esx-1 family proteins of the Type VII secretion systems of Mycobacterium bovis and Mycobacterium tuberculosis have been assessed and are frequently used as candidates for tuberculosis (TB) diagnosis in both humans and animals. The presence of ESAT-6 and CFP 10 proteins, which are the most immunogenic proteins of the Esx-1 system and have been widely investigated for the immunodiagnosis of tuberculosis, in some Mycobacteriaceae and in Mycobacterium leprae, poses limitations for their use in specific diagnoses of TB. As such, to improve the specificity of the ESAT-6/CFP 10-based cell-mediated immunity (CMI) assays, other proteins encoded by genes within and outside the RD 1 region of the esx-1 locus have been evaluated as candidate antigens for CMI, as well as to investigate humoral responses in combination with ESAT-6 and or CFP 10, with varying specificity and sensitivity results. Hence, in this study, we evaluated various non-tuberculous mycobacteria (NTM), Mycolicibacterium, Mycolicibacter and Mycobacteroides species genomes available on the NCBI database for the presence and composition of the RD1 region of the esx-1 locus. In addition, we also assayed by polymerase chain reaction (PCR) and sequencing of Mycobacteriaceae available in our culture collection for the presence and sequence diversity of esxA and esxB genes encoding ESAT-6 and CFP 10, respectively. Whole genome sequence (WGS) data analysis revealed the presence of RD 1 gene orthologs in 70 of the over 100 published genomes of pathogenic and non- pathogenic Mycobcteriaceae other than tuberculosis. Among species evaluated from our culture collection, in addition to earlier reports of the presence of esxA and esxB in certain Mycolicibacterium, Mycolicibacterium septicum/peregrinum, Mycolicibacterium porcinum and Mycobacterium sp. N845T were also found to harbour orthologs of both genes. Orthologs of esxA only were detected in Mycobacterium brasiliensis, Mycolicibacterium elephantis and Mycolicibacterium flouroantheinivorans, whereas in Mycolicibacter engbackii, Mycolicibacterium mageritense and Mycobacterium paraffinicum, only esxB orthologs were detected. A phylogenetic analysis based on esxA and esxB sequences separated slow-growing from rapidly growing bacteria. These findings strengthen previous suggestions that esxA and esxB may be encoded in the majority of Mycobacteriaceae. The role of the Esx-1 system in both pathogenic and non-pathogenic Mycobacteriaceae needs further investigation, as these species may pose limitations to immunological assays for TB.

There are different tuberculosis diagnostic tools available that detect an antigen-specific immune response. The present study aims to evaluate the potential of cross-reactive responses of a CFP-10 and ESAT-6 antigen-based TB test using bioinformatics tools. The study found that the presence of the sequences coding for the CFP-10 and ESAT-6 antigens in mycobacterial genomes is not associated with their pathogenicity, and not even consistent within a single species among its strains, which can lead to either false positive or false negative test results. The data that was analyzed included genome assemblies of all available mycobacterial strains obtained from the NCBI Genome database, while the standalone BLAST and tblastn programs were utilized to detect the presence of the CFP-10 and ESAT-6 sequences. The findings revealed that a number of non-pathogenic mycobacteria contained the aforementioned sequences, while some pathogenic mycobacteria did not, indicating that a standard tuberculin skin test should be more preferable for detecting various pathogenic mycobacteria compared to antigen-specific tests. In the Mycobacterium tuberculosis complex (MTBC), the proportion of positive strains varied within individual species, indicating a complex relationship. Among non-tuberculous mycobacteria (NTMB), more than half of the analyzed species did not contain these sequences which is consistent with their non-pathogenicity. Further research is necessary to fully comprehend the relationship between MTBC pathogenicity and the CFP-10 and ESAT-6 sequences. This could lead to a conclusion that a standard tuberculin skin test, although non-specific due to the undefined antigen content, may be able to detect various pathogenic mycobacteria in a more reliable manner than antigen-specific tests.

The retina undergoes compensatory changes in response to progressive photoreceptor loss/dysfunction; however, studies of inherited retinal diseases (IRDs) often lack a temporal connection between gene expression and visual function. Here, we used three mouse models of IRD - Cnga3-/-, Pde6ccpfl1, and Rd1 - to investigate over time the effect of photoreceptor degeneration, particularly cones, on visual function and gene expression. Changes to gene expression include increases in cell survival and cell death genes in Pde6ccpfl1 before significant cell loss, as well as an increase in cone-specific genes in the Rd1 at the peak of rod death. We show that Cnga3-/- and Pde6ccpfl1 mice maintained photopic visual acuity via optomotor responses, despite no recordable cone electroretinogram (ERG), while functional measures and photoreceptors loss were correlated in Rd1 mice. There were also significant changes to oscillatory potentials (OPs) in Cnga3-/- and Pde6ccpfl1, implying an effect on inner retinal cells as a result of cone degeneration. These results indicate a potentially malleable retinal environment following cone degeneration; however, further investigation is needed to elucidate how these changes compensate for the loss of cone function.

Tuberculosis remains a global health threat with high morbidity. Dendritic cells (DCs) participate in the acute and chronic inflammatory responses to Mycobacterium tuberculosis (Mtb) by directing the adaptive immune response and are present in lung granulomas. In macrophages, the interaction of lipid droplets (LDs) with mycobacteria-containing phagosomes is central to host-pathogen interactions. However, the data available for DCs are still a matter of debate. Here, we reported that bone marrow-derived DCs (BMDCs) were susceptible to Mtb infection and replication at similar rate to macrophages. Unlike macrophages, the analysis of gene expression showed that Mtb infection induced a delayed increase in lipid droplet-related genes and proinflammatory response. Hence, LD accumulation has been observed by high-content imaging in late periods. Infection of BMDCs with killed H37Rv demonstrated that LD accumulation depends on Mtb viability. Moreover, infection with the attenuated strains H37Ra and Mycobacterium bovis-BCG induced only an early transient increase in LDs, whereas virulent Mtb also induced delayed LD accumulation. In addition, infection with the BCG strain with the reintroduced virulence RD1 locus induced higher LD accumulation and bacterial replication when compared to parental BCG. Collectively, our data suggest that delayed LD accumulation in DCs is dependent on mycobacterial viability and virulence.

Insulin-degrading enzyme (IDE) was named after its role as a proteolytic enzyme of insulin. However, recent findings suggest that IDE is a widely expressed, multitask protein, with both proteolytic and non-proteolytic functions. Here, we characterize the expression of IDE in the mammalian retina in both physiological and pathological conditions. We found that IDE was enriched in cone inner segments. IDE levels were downregulated in the dystrophic retina of several mouse models of retinitis pigmentosa carrying distinct mutations. In rd10 mice, a commonly studied mouse model of retinitis pigmentosa, treatment with an IDE activator (a synthetic peptide analog of preimplantation factor) delayed loss of visual function and preserved photoreceptor cells. Together, these results point to potential novel roles for IDE in retinal physiology and disease, further extending the list of diverse functions attributed to this enzyme.

Retinitis pigmentosa (RP) is an inherited retinal degenerative disease that begins with defective rod photoreceptor function, followed by impaired cone function, and complete blindness in its late stage. To date, however, there is no effective treatment for RP. By carrying a nonsense mutation in the Pde6b gene, rd1 mice display elevated cGMP in conjunction with higher intracellular Ca 2+ in their rod photoreceptors, resulting in fast retinal degeneration. Ca 2+ has been linked to activation of the mammalian target of rapamycin (mTOR) pathway. The mTOR pathway integrates extracellular and intracellular signals to sense the supply of nutrients and plays a central role in regulating protein and lipid synthesis as well as apoptosis and autophagy. In the present study, we showed that mTOR and phosphorylated mTOR (p-mTOR, activated form of mTOR) are up-regulated in rd1 photoreceptors at postnatal day 10 (P10), a pre-degenerative stage. Moreover, the downstream effectors of mTOR, such as pS6K and S6K, are also increased, suggesting activation of the mTOR signaling pathway. Intravitreal administration of rapamycin, a negative regulator of mTOR, inhibits the mTOR pathway in rd1 photoreceptors. Consequently, the progression of retinal degeneration is slower and retinal function is enhanced, possibly mediated by activation of autophagy in the photoreceptors. Taken together, these results highlight rapamycin as a potential therapeutic avenue for retinal degeneration.
视网膜色素变性（RP）是一种遗传性视网膜变性疾病。RP最初影响视杆细胞的功能，随着病情的发展，视锥细胞也会受损，最终导致完全失明。该病目前没有有效的治疗方法。 rd1小鼠是一种被广泛用于研究视网膜变性的小鼠模型。 rd1小鼠携带天然的 Pde6b基因无义突变，其视杆细胞缺乏有活性的PDE6酶，引起胞内cGMP水平升高，更多的Ca 2+通道打开，使大量的Ca 2+进入视杆细胞，导致视网膜感光细胞迅速发生变性。以前的研究表明，胞内高水平的Ca 2+会激活mTOR信号通路，该通路感知细胞内外养分的变化，调节蛋白和脂类物质的合成，并调控细胞的凋亡和自噬。在本研究中我们发现 rd1小鼠感光细胞中mTOR信号通路被激活。P10的 rd1小鼠（视网膜发生明显变性之前）经玻璃体腔注射mTOR的抑制剂雷帕霉素治疗后，视网膜变性显著延缓，视网膜功能得到显著改善。雷帕霉素的治疗作用可能通过促进细胞自噬实现。综上，雷帕霉素有望成为一种治疗视网膜变性的药物。.

Many RD-causing mutations lead to a dysregulation of cyclic guanosine monophosphate (cGMP), making cGMP signalling a prime target for the development of new treatment approaches. We showed previously that an analogue of cGMP, which inhibited cGMP signalling targets, increased photoreceptor viability in three rodent RD models carrying different genetic defects, in different RD genes. This raises the question of the possible generality of this approach as a treatment for RD. Here, we review RD genes that can be associated with high cGMP and discuss which RD genes might be amenable to a treatment aimed at inhibiting excessive cGMP signalling.

In humans cone photoreceptors are responsible for high-resolution colour vision. A variety of retinal diseases can compromise cone viability, and, at present, no satisfactory treatment options are available. Here, we present data towards establishing a reliable, high-throughput assay system that will facilitate the search for cone neuroprotective compounds using the murine-photoreceptor cell line 661 W. To further characterize 661 W cells, a retinal marker study was performed, followed by the induction of cell death using paradigms over-activating cGMP-dependent protein kinase G (PKG). We found that 661 W cells may be used to mimic specific aspects of cone degeneration and may thus be valuable for future compound screening studies.

Retinitis Pigmentosa (RP) is a class of hereditary retinal dystrophy associated with gradual visual failure and a subsequent loss of light-sensitive cells in the retina, leading to blindness. Many mutated genes were found to be causative of this disease. Despite a number of compiling efforts, the process of cell death in photoreceptors remains to be clearly elucidated. We recently reported an abnormal cell cycle reentry in photoreceptors undergoing degeneration in Rd1 mice, a model of RP, and identified the polycomb repressive complex 1 (PRC1) core component BMI1 as a critical molecular factor orchestrating the cell death mechanism. As the cell death rescue in Rd1;Bmi-1 KO mice was independent on the conventional Ink4a/Arf pathways, we now explored the structural properties of BMI1 in order to examine the differential expression of its posttranslational modifications in Rd1 retina. Our results suggest that BMI1 cell death induction in Rd1 is not related to its phosphorylation status. We therefore propose the epigenetic activity of BMI1 as an alternative route for BMI1-mediated toxicity in Rd1.

The 14-3-3 family of proteins has undergone considerable expansion in higher eukaryotes with humans and mice expressing seven isoforms (β, ε, η, γ, θ, ζ, and σ) from seven distinct genes (YWHAB, YWAHE, YWHAH, YWHAG, YWHAQ, YWHAZ, and SFN). Growing evidence indicates that while highly conserved, these isoforms are not entirely functionally redundant as they exhibit unique tissue expression profiles, subcellular localization, and biochemical functions. A key limitation in our understanding of 14-3-3 biology lies in our limited knowledge of cell-type specific 14-3-3 expression. Here we provide a characterization of 14-3-3 expression in whole retina and isolated rod photoreceptors using reverse-transcriptase digital droplet PCR. We find that all 14-3-3 genes with the exception of SFN are expressed in mouse retina with YWHAQ and YWHAE being the most highly expressed. Rod photoreceptors are enriched in YWHAE (14-3-3 ε). Immunohistochemistry revealed that 14-3-3 ε and 14-3-3 ζ exhibit unique distributions in photoreceptors with 14-3-3 ε restricted to the inner segment and 14-3-3 ζ localized to the outer segment. Our data demonstrates that, in the retina, 14-3-3 isoforms likely serve specific functions as they exhibit unique expression levels and cell-type specificity. As such, future investigations into 14-3-3 function in rod photoreceptors should be centered on 14-3-3 ε and 14-3-3 ζ, depending on the subcellular region of question.