Is there a prechordal region and an acroterminal domain in amphioxus?

This essay re-examines the singular case of the supposedly unique rostrally elongated notochord described classically in amphioxus. We start from our previous observations in hpf 21 larvae [Albuixech-Crespo et al., 2017] indicating that the brain vesicle has rostrally a rather standard hypothalamic molecular configuration. This correlates with the notochord across a possible rostromedian acroterminal hypothalamic domain . The notochord shows some molecular differences that specifically characterize its pre-acroterminal extension beyond its normal rostral end under the mamillary region. We explored an alternative interpretation that the putative extension of this notochord actually represents a variant form of the prechordal plate in amphioxus, some of whose cells would adopt the notochordal typology, but would lack notochordal patterning properties, and might have some (but not all) prechordal ones instead. We survey in detail the classic and recent literature on gastrulation, prechordal plate and notochord formation in amphioxus, compared the observed patterns with those of some other vertebrates of interest, and re-examine the literature on differential gene expression patterns in this rostralmost area of the head. We noted that previous literature failed at identifying the amphioxus prechordal primordia at appropriate stages. Under this interpretation, a consistent picture can be drawn for cephalochordates, tunicates, and vertebrates. Moreover, there is little evidence for an intrinsic capacity of the early notochord to grow rostralwards (it normally elongates caudalwards). Altogether, we conclude that the hypothesis of a prechordal nature of the elongated amphioxus notochord is consistent with the evidence presented.

Manganese Is a Strong Specific Activator of the RNA Synthetic Activity of Human Polη

DNA polymerase η (Polη) is a translesion synthesis polymerase that can bypass different DNA lesions with varying efficiency and fidelity. Its most well-known function is the error-free bypass of ultraviolet light-induced cyclobutane pyrimidine dimers. The lack of this unique ability in humans leads to the development of a cancer-predisposing disease, the variant form of xeroderma pigmentosum. Human Polη can insert rNTPs during DNA synthesis, though with much lower efficiency than dNTPs, and it can even extend an RNA chain with ribonucleotides. We have previously shown that Mn2+ is a specific activator of the RNA synthetic activity of yeast Polη that increases the efficiency of the reaction by several thousand-fold over Mg2+. In this study, our goal was to investigate the metal cofactor dependence of RNA synthesis by human Polη. We found that out of the investigated metal cations, only Mn2+ supported robust RNA synthesis. Steady state kinetic analysis showed that Mn2+ activated the reaction a thousand-fold compared to Mg2+, even during DNA damage bypass opposite 8-oxoG and TT dimer. Our results revealed a two order of magnitude higher affinity of human Polη towards ribonucleotides in the presence of Mn2+ compared to Mg2+. It is noteworthy that activation occurred without lowering the base selectivity of the enzyme on undamaged templates, whereas the fidelity decreased across a TT dimer. In summary, our data strongly suggest that, like with its yeast homolog, Mn2+ is the proper metal cofactor of hPolη during RNA chain extension, and selective metal cofactor utilization contributes to switching between its DNA and RNA synthetic activities.

Biosynthetic Mechanisms and Biological Significance of Glycerol Phosphate-Containing Glycan in Mammals

Bacteria contain glycerol phosphate (GroP)-containing glycans, which are important constituents of cell-surface glycopolymers such as the teichoic acids of Gram-positive bacterial cell walls. These glycopolymers comprising GroP play crucial roles in bacterial physiology and virulence. Recently, the first identification of a GroP-containing glycan in mammals was reported as a variant form of O-mannosyl glycan on α-dystroglycan (α-DG). However, the biological significance of such GroP modification remains largely unknown. In this review, we provide an overview of this new discovery of GroP-containing glycan in mammals and then outline the recent progress in elucidating the biosynthetic mechanisms of GroP-containing glycans on α-DG. In addition, we discuss the potential biological role of GroP modification along with the challenges and prospects for further research. The progress in this newly identified glycan modification will provide insights into the phylogenetic implications of glycan.

Genetic and physical interactions between Polη and Rev1 in response to UV-induced DNA damage in mammalian cells

In response to UV irradiation, translesion DNA synthesis (TLS) utilizes specialized DNA polymerases to bypass replication-blocking lesions. In a well-established polymerase switch model, Polη is thought to be a preferred TLS polymerase to insert correct nucleotides across from the thymine dimer, and Rev1 plays a scaffold role through physical interaction with Polη and the Rev7 subunit of Polζ for continual DNA synthesis. Defective Polη causes a variant form of xeroderma pigmentosum (XPV), a disease with predisposition to sunlight-induced skin cancer. Previous studies revealed that expression of Rev1 alone is sufficient to confer enhanced UV damage tolerance in mammalian cells, which depends on its physical interaction with Polζ but is independent of Polη, a conclusion that appears to contradict current literature on the critical roles of Polη in TLS. To test a hypothesis that the Rev1 catalytic activity is required to backup Polη in TLS, we found that the Rev1 polymerase-dead mutation is synergistic with either Polη mutation or the Polη-interaction mutation in response to UV-induced DNA damage. On the other hand, functional complementation of polH cells by Polη relies on its physical interaction with Rev1. Hence, our studies reveal critical interactions between Rev1 and Polη in response to UV damage.

Non-painful severe variant form of eruptive lingual papillitis: A case report and literature review

Eruptive lingual papillitis is a common benign disorder manifested by inflammation of fungiform papillae on the dorsolateral surface of the tongue. Several variants of lingual papillitis have been reported since 1997, most or all of them with painful erythematous papules. Here we report a case of 6 years old girl child with non-painful severe variant form of eruptive lingual papillitis presented to the emergency department. The entire dorsal surface of the tongue was surfaced by 2-3mm by multiple erythematous papules and some with a white or yellowish colour. The papules were excessively inflamed, pigmented, aggregated, and crusted. The cause was idiopathic which resolved within ten days. The parent and patient were reassured with advice to practice oral hygiene. This is a rare case report describing non-painful lingual papillitis without a history of any prior episodes.

A Note on Sufi Snakes and Ladders

Among the diverse treasures of the Royal Asiatic Society are two early nineteenth century paper playing boards for the north Indian game of gyān caupar̩, the ‘Chaupar of Knowledge (Gnosis)’. This once popular game, played with dice or cowry shells, leads its players gradually up the board from hellish states or earthly vices to higher virtues and ultimately to heaven or liberation. It is known in various Jain and Hindu (mainly Vaiṣṇava) versions of the eighteenth to nineteenth centuries and a small handful of nineteenth century Muslim (or Sufi) examples. By the 1890s it also gave rise, in a simplified and denatured form, to the English children's game of Snakes and Ladders. One of the Society's boards is an ingenious 124-square version of the Vaiṣṇava form of gyān caupar̩, unique in its design and philosophical conception, whose inventor has been identified as the Brahmin scholar Thiruvenkatacharya Shastri. The Society's other board is a rare example of the 100-square Muslim form of gyān caupar̩ (Fig. 1), inscribed with Persian and Arabic square names that are loosely based around Sufi terms for the stages of the mystical path. I am here concerned with this form of the game and in particular with an expanded variant form of it that has recently come to light.

A Single Nucleotide Polymorphism in lptG Increases Tolerance to Bile Salts, Acid, and Staining of Calcofluor-Binding Polysaccharides in Salmonella enterica Serovar Typhimurium E40

The outer membrane of Salmonella enterica plays an important role in combating stress encountered in the environment and hosts. The transport and insertion of lipopolysaccharides (LPS) into the outer membrane involves lipopolysaccharide transport proteins (LptA-F) and mutations in the genes encoding for these proteins are often lethal or result in the transport of atypical LPS that can alter stress tolerance in bacteria. During studies of heterogeneity in bile salts tolerance, S. enterica serovar Typhimurium E40 was segregated into bile salts tolerant and sensitive cells by screening for growth in TSB with 10% bile salts. An isolate (E40V) with a bile salts MIC >20% was selected for further characterization. Whole-genome sequencing of E40 and E40V using Illumina and PacBio SMRT technologies revealed a non-synonymous single nucleotide polymorphism (SNP) in lptG. Leucine at residue 26 in E40 was substituted with proline in E40V. In addition to growth in the presence of 10% bile salts, E40V was susceptible to novobiocin while E40 was not. Transcriptional analysis of E40 and E40V, in the absence of bile salts, revealed significantly greater (p < 0.05) levels of transcript in three genes in E40V; yjbE (encoding for an extracellular polymeric substance production protein), yciE (encoding for a putative stress response protein), and an uncharacterized gene annotated as an acid shock protein precursor (ASPP). No transcripts of genes were present at a greater level in E40 compared to E40V. Corresponding with the greater level of these transcripts, E40V had greater survival at pH 3.35 and staining of Calcofluor-binding polysaccharide (CBPS). To confirm the SNP in lptG was associated with these phenotypes, strain E40E was engineered from E40 to encode for the variant form of LptG (L26P). E40E exhibited the same differences in gene transcripts and phenotypes as E40V, including susceptibility to novobiocin, confirming the SNP was responsible for these differences.

#31: Children with Invasive S. aureus Infection Produce Broadly Neutralizing Antibodies Against Distantly Related Variants of the Cytotoxin LukAB

Background Staphylococcus aureus is the most common invasive bacterial pathogen in children, and novel targets of intervention are urgently needed. The two-component leukotoxin, LukAB, is critical for S. aureus targeting and killing of human neutrophils and is abundantly produced in the setting of invasive human infection. LukAB is unique among S. aureus cytotoxins in that it exists in variant form across distantly related strains of clinically relevant S. aureus. The broad diversity of circulating clinical S. aureus strains must be taken into account to successfully develop new anti-staphylococcal preventives or therapeutics. This diversity and ongoing S. aureus evolution may explain previous unsuccessful attempts to intervene against S. aureus at the virulence factor level. We have previously shown that LukAB is ubiquitous among clinical isolates and that children with invasive disease mount a high-titer neutralizing response, but the breadth of function of this response has not previously been explored. Methods B-cells were isolated from children with invasive S. aureus infections (e.g. bacteremia or acute hematogenous osteomyelitis). Following EBV-transformation, cell supernatants were screened for LukAB binding and selected for generation of monoclonal hybridomas. MAbs were assessed for binding by ELISA and neutralizing function by in vitro cytotoxicity assays, where neutrophil-like HL-60 cells were cultured in the presence of human mAbs and diverse allelic variants of LukAB. The infecting S. aureus isolates were typed using multilocus sequence typing (MLST) to determine clonal complexes. Results 34 distinct human anti-LukAB mAbs were generated from 3 children with invasive S. aureus disease. Of these, 22% were isolated following infection with a strain belonging to clonal complex 8 (CC8), consistent with the epidemic USA300 clone, and 78% were generated against CC5 strains. Within this panel, all mAbs potently neutralized LukAB from the same clonal complex as the corresponding infecting isolate, but 23 also demonstrated neutralization against other allelic variants of LukAB. 7 mAbs were capable of broad, potent neutralization against LukAB variants from all clinically relevant clonal complexes tested (CC8, CC30, CC45, CC75, CC1, CC5, and CC398). Conclusions A subset of human mAbs isolated from children with invasive S. aureus disease were capable of broad neutralization against distantly related variants of the important toxin LukAB. This has two important implications: First, we found strong evidence of a conserved target (or targets) for antibody-mediated toxin neutralization across diverse strains of S. aureus. Second, this provides additional support for this toxin as a target of intervention, as some previous vaccine attempts were likely unsuccessful due to activity against a narrow subset of circulating S. aureus strains.

In Cellulo Examination of a Beta-Alpha Hybrid Construct of Beta-Hexosaminidase A Subunits, Reported to Interact with the GM2 Activator Protein and Hydrolyze GM2 Ganglioside

The hydrolysis in lysosomes of GM2 ganglioside to GM3 ganglioside requires the correct synthesis, intracellular assembly and transport of three separate gene products; i.e., the alpha and beta subunits of heterodimeric beta-hexosaminidase A, E.C. # 3.2.1.52 (encoded by the HEXA and HEXB genes, respectively), and the GM2-activator protein (GM2AP, encoded by the GM2A gene). Mutations in any one of these genes can result in one of three neurodegenerative diseases collectively known as GM2 gangliosidosis (HEXA, Tay-Sachs disease, MIM # 272800; HEXB, Sandhoff disease, MIM # 268800; and GM2A, AB-variant form, MIM # 272750). Elements of both of the hexosaminidase A subunits are needed to productively interact with the GM2 ganglioside-GM2AP complex in the lysosome. Some of these elements have been predicted from the crystal structures of hexosaminidase and the activator. Recently a hybrid of the two subunits has been constructed and reported to be capable of forming homodimers that can perform this reaction in vivo, which could greatly simplify vector-mediated gene transfer approaches for Tay-Sachs or Sandhoff diseases. A cDNA encoding a hybrid hexosaminidase subunit capable of dimerizing and hydrolyzing GM2 ganglioside could be incorporated into a single vector, whereas packaging both subunits of hexosaminidase A into vectors, such as adeno-associated virus, would be impractical due to size constraints. In this report we examine the previously published hybrid construct (H1) and a new more extensive hybrid (H2), with our documented in cellulo (live cell- based) assay utilizing a fluorescent GM2 ganglioside derivative. Unfortunately when Tay-Sachs cells were transfected with either the H1 or H2 hybrid construct and then were fed the GM2 derivative, no significant increase in its turnover was detected. In vitro assays with the isolated H1 or H2 homodimers confirmed that neither was capable of human GM2AP-dependent hydrolysis of GM2 ganglioside.

In Cellulo Examination of a Beta-Alpha Hybrid Construct of Beta-Hexosaminidase A Subunits, Reported to Interact with the GM2 Activator Protein and Hydrolyze GM2 Ganglioside

The hydrolysis in lysosomes of GM2 ganglioside to GM3 ganglioside requires the correct synthesis, intracellular assembly and transport of three separate gene products; i.e., the alpha and beta subunits of heterodimeric beta-hexosaminidase A, E.C. # 3.2.1.52 (encoded by the HEXA and HEXB genes, respectively), and the GM2-activator protein (GM2AP, encoded by the GM2A gene). Mutations in any one of these genes can result in one of three neurodegenerative diseases collectively known as GM2 gangliosidosis (HEXA, Tay-Sachs disease, MIM # 272800; HEXB, Sandhoff disease, MIM # 268800; and GM2A, AB-variant form, MIM # 272750). Elements of both of the hexosaminidase A subunits are needed to productively interact with the GM2 ganglioside-GM2AP complex in the lysosome. Some of these elements have been predicted from the crystal structures of hexosaminidase and the activator. Recently a hybrid of the two subunits has been constructed and reported to be capable of forming homodimers that can perform this reaction in vivo, which could greatly simplify vector-mediated gene transfer approaches for Tay-Sachs or Sandhoff diseases. A cDNA encoding a hybrid hexosaminidase subunit capable of dimerizing and hydrolyzing GM2 ganglioside could be incorporated into a single vector, whereas packaging both subunits of hexosaminidase A into vectors, such as adeno-associated virus, would be impractical due to size constraints. In this report we examine the previously published hybrid construct (H1) and a new more extensive hybrid (H2), with our documented in cellulo (live cell- based) assay utilizing a fluorescent GM2 ganglioside derivative. Unfortunately when Tay-Sachs cells were transfected with either the H1 or H2 hybrid construct and then were fed the GM2 derivative, no significant increase in its turnover was detected. In vitro assays with the isolated H1 or H2 homodimers confirmed that neither was capable of human GM2AP-dependent hydrolysis of GM2 ganglioside.