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ADAMTS13, a metalloproteinase, specifically cleaves unusually large multimers of von Willebrand factor (VWF), newly released from vascular endothelial cells. The ratio of ADAMTS13 activity to VWF antigen (ADAMTS13/VWF) and indicators of the alternative complement pathway (C3a and sC5b-9) are both related to the severity of COVID-19. The ADAMTS13/VWF ratio is generally moderately decreased (0.18–0.35) in patients with severe COVID-19. When these patients experience cytokine storms, both interleukin-8 and TNFα stimulate VWF release from vascular endothelial cells, while interleukin-6 inhibits both production of ADAMTS13 and its interaction with VWF, resulting in localized severe deficiency of ADAMTS13 activity. Platelet factor 4 and thrombospondin-1, both released upon platelet activation, bind to the VWF-A2 domain and enhance the blockade of ADAMTS13 function. Thus, the released unusually-large VWF multimers remain associated with the vascular endothelial cell surface, via anchoring with syndecan-1 in the glycocalyx. Unfolding of the VWF-A2 domain, which has high sequence homology with complement factor B, allows the domain to bind to activated complement C3b, providing a platform for complement activation of the alternative pathway. The resultant C3a and C5a generate tissue factor-rich neutrophil extracellular traps (NETs), which induce the mixed immunothrombosis, fibrin clots and platelet aggregates typically seen in patients with severe COVID-19. [ABSTRACT FROM AUTHOR]

Over the past 200 years, almost every invertebrate phylum has been proposed as a starting point for evolving vertebrates. Most of these scenarios are outdated, but several are still seriously considered. The short-range transition from ancestral invertebrate chordates (similar to amphioxus and tunicates) to vertebrates is well accepted. However, longer-range transitions leading up to the invertebrate chordates themselves are more controversial. Opinion is divided between the annelid and the enteropneust scenarios, predicting, respectively, a complex or a simple ancestor for bilaterian animals. Deciding between these ideas will be facilitated by further comparative studies of multicellular animals, including enigmatic taxa such as xenacoelomorphs. [ABSTRACT FROM AUTHOR]

The vertebrate brain is highly complex with millions to billions of neurons. During development, the neural plate border region gives rise to the neural crest, cranial placodes and, in anamniotes, to Rohon-Beard sensory neurons, whereas the boundary region of the midbrain and hindbrain develops organizer properties. Comparisons of developmental gene expression and neuroanatomy between vertebrates and the basal chordate amphioxus, which has only thousands of neurons and lacks a neural crest, most placodes and a midbrain–hindbrain organizer, indicate that these vertebrate features were built on a foundation already present in the ancestral chordate. Recent advances in genomics have provided insights into the elaboration of the molecular toolkit at the invertebrate–vertebrate transition that may have facilitated the evolution of these vertebrate characteristics. [ABSTRACT FROM AUTHOR]

Cephalochordates are the basal invertebrate chordates within the phylum Chordata. They are widely used as a model system for research in evolutionary developmental biology (EvoDevo) to understand the basic patterning mechanisms for the chordate body plan and the origin of vertebrates. Recently, the genome of the cephalochordate Branchiostoma floridae was sequenced, which further brings this organism to the front for comparative genomic studies. In this paper, we report the generation of large-scale 5′- and 3′-expressed sequence tags (ESTs) from B. floridae and the complementary deoxyribonucleic acid (cDNA) resource for this species . Both 5′- and 3′-ESTs were sequenced for approximately 140,000 cDNA clones derived from five developmental stages, and the cDNA clones were subsequently grouped into independent clusters using 3′-EST sequences. We identified 21,229 cDNA clusters, and each corresponds to a unique transcript species from B. floridae. We then chose 24,020 cDNA clones representing all of these 21,229 clusters to generate the “ Branchiostoma floridae Gene Collection Release 1.” We also constructed a database with a searchable interface for this EST dataset and the related information on “ Branchiostoma floridae Gene Collection Release 1.” This set of cDNA clones along with our cDNA database will serve as an important resource for future research in this basal chordate. This Gene Collection and the original 140,000 individual cDNA clones are available to the research community upon request. [ABSTRACT FROM AUTHOR]

In the basal chordate amphioxus ( Branchiostoma), somites extend the full length of the body. The anteriormost somites segment during the gastrula and neurula stages from dorsolateral grooves of the archenteron. The remaining ones pinch off, one at a time, from the tail bud. These posterior somites appear to be homologous to those of vertebrates, even though the latter pinch off from the anterior end of bands of presomitic mesoderm rather than directly from the tail bud. To gain insights into the evolution of mesodermal segmentation in chordates, we determined the expression of ten genes in nascent amphioxus somites. Five ( Uncx4. 1, NeuroD/ atonal- related, IrxA, Pcdhδ2- 17/ 18, and Hey1) are expressed in stripes in the dorsolateral mesoderm at the gastrula stage and in the tail bud while three ( Paraxis, Lcx, and Axin) are expressed in the posterior mesendoderm at the gastrula and neurula stages and in the tail bud at later stages. Expression of two genes ( Pbx and OligA) suggests roles in the anterior somites that may be unrelated to initial segmentation. Together with previous data, our results indicate that, with the exception that Engrailed is only segmentally expressed in the anterior somites, the genetic mechanisms controlling formation of both the anterior and posterior somites are probably largely identical. Thus, the fundamental pathways for mesodermal segmentation involving Notch–Delta, Wnt/β-catenin, and Fgf signaling were already in place in the common ancestor of amphioxus and vertebrates although budding of somites from bands of presomitic mesoderm exhibiting waves of expression of Notch, Wnt, and Fgf target genes was likely a vertebrate novelty. Given the conservation of segmentation gene expression between amphioxus and vertebrate somites, we propose that the clock mechanism may have been established in the basal chordate, while the wavefront evolved later in the vertebrate lineage. [ABSTRACT FROM AUTHOR]

Pax genes encode transcription factors critical for metazoan development. Large-scale gene duplication with subsequent gene losses during vertebrate evolution has resulted in two human genes for each of the Pax1/9, Pax3/7, and Pax4/6 subfamilies and three for the Pax2/5/8 subfamily, compared to one each in the cephalochordate amphioxus. In addition, alternative splicing occurs in vertebrate Pax transcripts from all four subfamilies, and many splice forms are known to have functional importance. To better understand the evolution of alternative splicing within the Pax family, we systematically surveyed transcripts of the four amphioxus Pax genes. We have found alternative splicing in every gene. Comparisons with vertebrates suggest that the number of alternative splicing events per gene has not decreased following duplication; there are comparable levels in the four amphioxus Pax genes as in each gene of the equivalent vertebrate families. Thus, the total number of isoforms for the nine vertebrate genes is considerably higher than for the four amphioxus genes. Most alternative splicing events appear to have arisen since the divergence of amphioxus and vertebrate lineages, suggesting that differences in alternative splicing could account for divergent functions of the highly conserved Pax genes in both lineages. However, several events predicted to dramatically alter known functional domains are conserved between amphioxus and vertebrates, suggestive of a common chordate function. Our results, together with previous studies of vertebrate Pax genes, support the theory that alternative splicing impacts functional motifs more than gene duplication followed by divergence. [ABSTRACT FROM AUTHOR]

The organizer of the vertebrate gastrula is an important signalling centre that induces and patterns dorsal axial structures. Although a topic of long-standing interest, the evolutionary origin of the organizer remains unclear. Here we show that the gastrula of the cephalochordate amphioxus expresses dorsal/ventral (D/V) patterning genes (for example, bone morphogenetic proteins (BMPs), Nodal and their antagonists) in patterns reminiscent of those of their vertebrate orthlogues, and that amphioxus embryos, like those of vertebrates, are ventralized by exogenous BMP protein. In addition, Wnt-antagonists (for example, Dkks and sFRP2-like) are expressed anteriorly, whereas Wnt genes themselves are expressed posteriorly, consistent with a role for Wnt signalling in anterior/posterior (A/P) patterning. These results suggest evolutionary conservation of the mechanisms for both D/V and A/P patterning of the early gastrula. In light of recent phylogenetic analyses placing cephalochordates basally in the chordate lineage, we propose that separate signalling centres for patterning the D/V and A/P axes may be an ancestral chordate character. [ABSTRACT FROM AUTHOR]

A full-length FoxQ-related gene (AmphiFoxQ2) was isolated from amphioxus. Expression is first detectable in the animal/anterior hemisphere at the mid blastula stage. The midpoint of this expression domain coincides with the anterior pole of the embryo and is offset dorsally by about 20° from the animal pole. During the gastrula stage, expression is limited to the anterior ectoderm. By the early neurula stage, expression remains in the anterior ectoderm and also appears in the adjacent anterior mesendoderm. By the early larval stages, expression is detectable in the anteriormost ectoderm and in the rostral tip of the notochord. AmphiFoxQ2 is never expressed anywhere except at the anterior tip of amphioxus embryos and larvae. This is the first gene known that exclusively marks the anterior pole of chordate embryos. It may, therefore, play an important role in establishing and/or maintaining the anterior/posterior axis. [ABSTRACT FROM AUTHOR]

In vertebrates, the orphan nuclear receptors of the COUP-TF group function as negative transcriptional regulators that inhibit the hormonal induction of target genes mediated by classical members of the nuclear hormone superfamily, such as the retinoic acid receptors (RARs) or the thyroid hormone receptors (TRs). To investigate the evolutionary conservation of the roles of COUP-TF receptors as negative regulators in the retinoid and thyroid hormone pathways, we have characterized AmphiCOUP-TF, the homologue of COUP-TFI and COUP-TFII, in the chordate amphioxus (Branchiostoma floridae), the closest living invertebrate relative of the vertebrates. Electrophoretic mobility shift assays (EMSA) showed that AmphiCOUP-TF binds to a wide variety of response elements, as do its vertebrate homologues. Furthermore, AmphiCOUP-TF is a transcriptional repressor that strongly inhibits retinoic acid-mediated transactivation. In situ hybridizations revealed expression of AmphiCOUP-TF in the nerve cord of late larvae, in a region corresponding to hindbrain and probably anterior spinal cord. Although the amphioxus nerve cord appears unsegmented at the gross anatomical level, this pattern reflects segmentation at the cellular level with stripes of expressing cells occurring adjacent to the ends and the centers of each myotomal segment, which may include visceral motor neurons and somatic motor neurons respectively, among other cells. A comparison of the expression pattern of AmphiCOUP-TF with those of its vertebrate homologues, suggests that the roles of COUP-TF in patterning of the nerve cord evolved prior to the split between the amphioxus and vertebrate lineages. Furthermore, in vitro data also suggest that Amphi-COUP-TF acts as a negative regulator of signalling by other nuclear receptors such as RAR, TR or ER. [ABSTRACT FROM AUTHOR]

We characterized an amphioxus NK-2 homeobox gene ( AmphiNk2–1), a homologue of vertebrate Nkx2–1, which is involved in the development of the central nervous system and thyroid gland. At the early neurula stage of amphioxus, AmphiNk2–1 expression is first detected medially in the neural plate. By the mid-neurula stage, expression is localized ventrally in the nerve cord and also begins in the endoderm. During the late neurula stage, the ventral neural expression becomes transiently segmented posteriorly and is then down-regulated except in the cerebral vesicle at the anterior end of the central nervous system. Within the cerebral vesicle AmphiNk2–1 is expressed in a broad ventral domain, probably comprising both the floor plate and basal plate regions; this pattern is comparable to Nkx2–1 expression in the mouse diencephalon. In the anterior part of the gut, expression becomes intense in the endostyle (the right wall of the pharynx), which is the presumed homologue of the vertebrate thyroid gland. More posteriorly, there is transitory expression in the midgut and hindgut. In sum, the present results help to support homologies (1) between the amphioxus endostyle and the vertebrate thyroid gland and (2) between the amphioxus cerebral vesicle and the vertebrate diencephalic forebrain. [ABSTRACT FROM AUTHOR]

The genome of amphioxus includes AmphiNk2-2, the first gene of the NK2 homeobox class to be demonstrated in any invertebrate deuterostome. AmphiNk2-2 encodes a protein with a TN domain, homeodomain, and NK2-specific domain; on the basis of amino acid identities in these conserved regions, AmphiNk2-2 is a homolog of Drosophila vnd and vertebrate Nkx2–2. During amphioxus development, expression of Amph- iNk2-2 is first detected ventrally in the endoderm of late gastrulae. In neurulae, endodermal expression divides into three domains (the pharynx, midgut, and hindgut), and neural expression commences in two longitudinal bands of cells in the anterior neural tube. These neural tube cells occupy a ventrolateral position on either side of the cerebral vesicle (the probable homolog of the vertebrate diencephalic forebrain). The dynamic expression patterns of AmphiNkx2-2 suggest successive roles, first in regionalizing the endoderm and nervous system and later during differentiation of specific cell types in the gut (possibly peptide endocrine cells) and brain (possibly including axon outgrowth and guidance). [ABSTRACT FROM AUTHOR]

The cephalochordate amphioxus is thought to share a common ancestor with vertebrates. To investigate the evolution of developmental mechanisms in chordates, cDNA clones for two amphioxus actin genes, BfCA1 and BfMA1, were isolated. BfCA1 encodes a cytoplasmic actin and is expressed in a variety of tissues during embryogenesis, beginning in the dorsolateral mesendoderm of the mid-gastrula. At the open neural plate stage, BfCA1 transcripts accumulate at the bases of the neuroectodermal cells adjacent the presumptive notochord. The 3’ untranslated region of BfCA1 contains a sequence that is similar to the ”zipcode” sequence of chicken β-cytoplasmic actin gene, which is thought to direct intracellular mRNA localization. BfCA1 is also expressed in the notochord through the early larval stage, in the pharynx and in the somites at the onset of muscle-cell differentiation. BfMA1 is a vertebrate-type muscle actin gene, although the deduced amino acid sequence is fairly divergent. Transcripts first appear in the early neurula in the somites as they begin to differentiate into axial muscle cells and persist into the adult stage. In young adults, transcripts are localized in the Z-discs of the muscle cells. Smooth muscle cells around the gill slits and striated muscle cells in the pterygeal muscle also express BfMA1; however, there is never any detectable expression in the notochord, which is a modified striated muscle. Together with the alkali myosin light chain gene AmphiMLC-alk, the sequence and muscle-specific expression of BfMA1 implies a conserved mechanism of muscle cell differentiation between amphioxus and vertebrates. Evolution of the chordate actin gene family is discussed based on molecular phylogenetic analysis and expression patterns of amphioxus actin genes. [ABSTRACT FROM AUTHOR]

The article focuses on a molecular study of the life cycle of tunicate. The genetic programs for patterning the early embryonic cycle are found to reveal a radical change during the development without complete disturbance of basic chordate body plan. However, while fundamental chordate body plan has retained, mechanism of retinoic-acid signaling that operates during such development has been lost. Such loss has prevented similar changes in other chordates.

A full-length Wnt1 gene (AmphiWnt1) was isolated from amphioxus. Expression is first detectable in the gastrula around the lip of the blastopore. By the early neurula, transcription is in the mesendoderm near the closed blastopore, but is down-regulated in the overlying ectoderm. In the late neurula, expression is limited to the posterior wall of the neurenteric canal. Later in development, AmphiWnt1 transcripts can no longer be detected. AmphiWnt1 has no counterpart of the predominant expression domains of vertebrate Wnt1 genes in the neural tube, but its expression may be more comparable to that of wingless in the invaginating hindgut primordium of insects. [ABSTRACT FROM AUTHOR]