ASU Scholarship Showcase

Neural progenitor cells (NPCs) derived from human pluripotent stem cells (hPSCs) are a multipotent cell population that is capable of nearly indefinite expansion and subsequent differentiation into the various neuronal and supporting cell types that comprise the CNS. However, current protocols for differentiating NPCs toward neuronal lineages result in a mixture of neurons from various regions of the CNS. In this study, we determined that endogenous WNT signaling is a primary contributor to the heterogeneity observed in NPC cultures and neuronal differentiation. Furthermore, exogenous manipulation of WNT signaling during neural differentiation, through either activation or inhibition, reduces this heterogeneity in NPC cultures, thereby promoting the formation of regionally homogeneous NPC and neuronal cultures. The ability to manipulate WNT signaling to generate regionally specific NPCs and neurons will be useful for studying human neural development and will greatly enhance the translational potential of hPSCs for neural-related therapies.

The threat of West Nile virus (WNV) epidemics with increasingly severe neuroinvasive infections demands the development and licensing of effective vaccines. To date, vaccine candidates based on inactivated, live-attenuated, or chimeric virus, and viral DNA and WNV protein subunits have been developed. Some have been approved for veterinary use or are under clinical investigation, yet no vaccine has been licensed for human use. Reaching the milestone of a commercialized human vaccine, however, may largely depend on the economics of vaccine production. Analysis suggests that currently only novel low-cost production technologies would allow vaccination to outcompete the cost of surveillance and clinical treatment. Here, we review progress using plants to address the economic challenges of WNV vaccine production. The advantages of plants as hosts for vaccine production in cost, speed and scalability, especially those of viral vector-based transient expression systems, are discussed. The progress in developing WNV subunit vaccines in plants is reviewed within the context of their expression, characterization, downstream processing, and immunogenicity in animal models. The development of vaccines based on enveloped and non-enveloped virus-like particles is also discussed. These advancements suggest that plants may provide a production platform that offers potent, safe and affordable human vaccines against WNV.

Although the majority of late-onset Alzheimer's disease (AD) patients are labeled sporadic, multiple genetic risk variants have been identified, the most powerful and prevalent of which is the e4 variant of the Apolipoprotein E (APOE) gene. Here, we generated human induced pluripotent stem cell (hiPSC) lines from the peripheral blood mononuclear cells (PBMCs) of a clinically diagnosed AD patient [ASUi003-A] and a non-demented control (NDC) patient [ASUi004-A] homozygous for the APOE4 risk allele. These hiPSCs maintained their original genotype, expressed pluripotency markers, exhibited a normal karyotype, and retained the ability to differentiate into cells representative of the three germ layers.

Nonsense-mediated RNA decay (NMD) is a highly conserved pathway that selectively degrades specific subsets of RNA transcripts. Here, we provide evidence that NMD regulates early human developmental cell fate. We found that NMD factors tend to be expressed at higher levels in human pluripotent cells than in differentiated cells, raising the possibility that NMD must be downregulated to permit differentiation. Loss- and gain-of-function experiments in human embryonic stem cells (hESCs) demonstrated that, indeed, NMD downregulation is essential for efficient generation of definitive endoderm. RNA-seq analysis identified NMD target transcripts induced when NMD is suppressed in hESCs, including many encoding signaling components. This led us to test the role of TGF-β and BMP signaling, which we found NMD acts through to influence definitive endoderm versus mesoderm fate. Our results suggest that selective RNA decay is critical for specifying the developmental fate of specific human embryonic cell lineages.

Previously, our group engineered a plant-derived monoclonal antibody (MAb) (pHu-E16) that efficiently treated West Nile virus (WNV) infection in mice. In this study, we developed several pHu-E16 variants to improve its efficacy. These variants included a single-chain variable fragment (scFv) of pHu-E16 fused to the heavy chain (HC) constant domains (CH1-3) of human IgG (pHu-E16scFv-CH1-3) and a tetravalent molecule (Tetra pHu-E16) assembled from pHu-E16scFv-CH1-3 with a second pHu-E16scFv fused to the light chain (LC) constant region. pHu-E16scFv-CH1-3 and Tetra pHu-E16 were efficiently expressed and assembled in plants. To assess the impact of differences in N-linked glycosylation on pHu-E16 variant assembly and function, we expressed additional pHu-E16 variants with various combinations of HC and LC components.

Our study revealed that proper pairing of HC and LC was essential for the complete N-glycan processing of antibodies in both plant and animal cells. Associated with their distinct N-glycoforms, pHu-E16, pHu-E16scFv-CH1-3 and Tetra pHu-E16 exhibited differential binding to C1q and specific Fcγ receptors (FcγR). Notably, none of the plant-derived Hu-E16 variants showed antibody-dependent enhancement (ADE) activity in CD32A+ human cells, suggesting the potential of plant-produced antibodies to minimize the adverse effect of ADE. Importantly, all plant-derived MAb variants exhibited at least equivalent in vitro neutralization and in vivo protection in mice compared to mammalian cell-produced Hu-E16. This study demonstrates the capacity of plants to express and assemble a large, complex and functional IgG-like tetravalent mAb variant and also provides insight into the relationship between MAb N-glycosylation, FcγR and C1q binding, and ADE. These new insights may allow the development of safer and cost effective MAb-based therapeutics for flaviviruses, and possibly other pathogens.

In the decade since Yamanaka and colleagues described methods to reprogram somatic cells into a pluripotent state, human induced pluripotent stem cells (hiPSCs) have demonstrated tremendous promise in numerous disease modeling, drug discovery, and regenerative medicine applications. More recently, the development and refinement of advanced gene transduction and editing technologies have further accelerated the potential of hiPSCs. In this review, we discuss the various gene editing technologies that are being implemented with hiPSCs. Specifically, we describe the emergence of technologies including zinc-finger nuclease (ZFN), transcription activator-like effector nuclease (TALEN), and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 that can be used to edit the genome at precise locations, and discuss the strengths and weaknesses of each of these technologies. In addition, we present the current applications of these technologies in elucidating the mechanisms of human development and disease, developing novel and effective therapeutic molecules, and engineering cell-based therapies. Finally, we discuss the emerging technological advances in targeted gene editing methods.

The Quadrangles Av-11 and Av-12 on Vesta are located at the northern rim of the giant Rheasilvia south polar impact basin. The primary geologic units in Av-11 and Av-12 include material from the Rheasilvia impact basin formation, smooth material and different types of impact crater structures (such as bimodal craters, dark and bright crater ray material and dark ejecta material). Av-11 and Av-12 exhibit almost the full range of mass wasting features observed on Vesta, such as slump blocks, spur-and-gully morphologies and landslides within craters. Processes of collapse, slope instability and seismically triggered events force material to slump down crater walls or scarps and produce landslides or rotational slump blocks. The spur-and-gully morphology that is known to form on Mars is also observed on Vesta; however, on Vesta this morphology formed under dry conditions.

A variety of geologic landforms and features are observed within quadrangle Av-13 Tuccia in the southern hemisphere of Vesta. The quadrangle covers parts of the highland Vestalia Terra as well as the floors of the large Rheasilvia and Veneneia impact basins, which results in a substantial elevation difference of more than 40 km between the northern and the southern portions of the quadrangle. Measurements of crater size–frequency distributions within and surrounding the Rheasilvia basin indicate that gravity-driven mass wasting in the interior of the basin has been important, and that the basin has a more ancient formation age than would be expected from the crater density on the basin floor alone. Subsequent to its formation, Rheasilvia was superimposed by several mid-sized impact craters. The most prominent craters are Tuccia, Eusebia, Vibidia, Galeria, and Antonia, whose geology and formation ages are investigated in detail in this work. These impact structures provide a variety of morphologies indicating different sorts of subsequent impact-related or gravity-driven mass wasting processes. Understanding the geologic history of the relatively young craters in the Rheasilvia basin is important in order to understand the even more degraded craters in other regions of Vesta.

In this paper we present a time-stratigraphic scheme and geologic time scale for the protoplanet Vesta, based on global geologic mapping and other analyses of NASA Dawn spacecraft data, complemented by insights gained from laboratory studies of howardite–eucrite–diogenite (HED) meteorites and geophysical modeling. On the basis of prominent impact structures and their associated deposits, we propose a time scale for Vesta that consists of four geologic time periods: Pre-Veneneian, Veneneian, Rheasilvian, and Marcian. The Pre-Veneneian Period covers the time from the formation of Vesta up to the Veneneia impact event, from 4.6 Ga to >2.1 Ga (using the asteroid flux-derived chronology system) or from 4.6 Ga to 3.7 Ga (under the lunar-derived chronology system). The Veneneian Period covers the time span between the Veneneia and Rheasilvia impact events, from >2.1 to 1 Ga (asteroid flux-derived chronology) or from 3.7 to 3.5 Ga (lunar-derived chronology), respectively. The Rheasilvian Period covers the time span between the Rheasilvia and Marcia impact events, and the Marcian Period covers the time between the Marcia impact event until the present. The age of the Marcia impact is still uncertain, but our current best estimates from crater counts of the ejecta blanket suggest an age between ∼120 and 390 Ma, depending upon choice of chronology system used. Regardless, the Marcia impact represents the youngest major geologic event on Vesta. Our proposed four-period geologic time scale for Vesta is, to a first order, comparable to those developed for other airless terrestrial bodies.

We report on a preliminary global geologic map of Vesta, based on data from the Dawn spacecraft’s High-Altitude Mapping Orbit (HAMO) and informed by Low-Altitude Mapping Orbit (LAMO) data. This map is part of an iterative mapping effort; the geologic map has been refined with each improvement in resolution. Vesta has a heavily-cratered surface, with large craters evident in numerous locations. The south pole is dominated by an impact structure identified before Dawn’s arrival. Two large impact structures have been resolved: the younger, larger Rheasilvia structure, and the older, more degraded Veneneia structure. The surface is also characterized by a system of deep, globe-girdling equatorial troughs and ridges, as well as an older system of troughs and ridges to the north. Troughs and ridges are also evident cutting across, and spiraling arcuately from, the Rheasilvia central mound.

However, no volcanic features have been unequivocally identified. Vesta can be divided very broadly into three terrains: heavily-cratered terrain; ridge-and-trough terrain (equatorial and northern); and terrain associated with the Rheasilvia crater. Localized features include bright and dark material and ejecta (some defined specifically by color); lobate deposits; and mass-wasting materials. No obvious volcanic features are evident. Stratigraphy of Vesta’s geologic units suggests a history in which formation of a primary crust was followed by the formation of impact craters, including Veneneia and the associated Saturnalia Fossae unit. Formation of Rheasilvia followed, along with associated structural deformation that shaped the Divalia Fossae ridge-and-trough unit at the equator. Subsequent impacts and mass wasting events subdued impact craters, rims and portions of ridge-and-trough sets, and formed slumps and landslides, especially within crater floors and along crater rims and scarps. Subsequent to the formation of Rheasilvia, discontinuous low-albedo deposits formed or were emplaced; these lie stratigraphically above the equatorial ridges that likely were formed by Rheasilvia. The last features to be formed were craters with bright rays and other surface mantling deposits.

Executed progressively throughout data acquisition, the iterative mapping process provided the team with geologic proto-units in a timely manner. However, interpretation of the resulting map was hampered by the necessity to provide the team with a standard nomenclature and symbology early in the process. With regard to mapping and interpreting units, the mapping process was hindered by the lack of calibrated mineralogic information. Topography and shadow played an important role in discriminating features and terrains, especially in the early stages of data acquisition.