Spontaneous travelling cortical waves gate perception in behaving primates.

Perceptual sensitivity varies from moment to moment. One potential source of this variability is spontaneous fluctuations in cortical activity that can travel as waves1. Spontaneous travelling waves have been reported during anaesthesia2-7, but it is not known whether they have a role during waking perception. Here, using newly developed analytic techniques to characterize the moment-to-moment dynamics of noisy multielectrode data, we identify spontaneous waves of activity in the extrastriate visual cortex of awake, behaving marmosets (Callithrix jacchus). In monkeys trained to detect faint visual targets, the timing and position of spontaneous travelling waves before target onset predicted the magnitude of target-evoked activity and the likelihood of target detection. By contrast, spatially disorganized fluctuations of neural activity were much less predictive. These results reveal an important role for spontaneous travelling waves in sensory processing through the modulation of neural and perceptual sensitivity.

Safety and Virologic Impact of Haploidentical NK Cells Plus Interleukin 2 or N-803 in HIV Infection.

BACKGROUND: Natural killer (NK) cells are dysfunctional in chronic human immunodeficiency virus (HIV) infection as they are not able to clear virus. We hypothesized that an infusion of NK cells, supported by interleukin 2 (IL-2) or IL-15, could decrease virus-producing cells in the lymphatic tissues. METHODS: We conducted a phase 1 pilot study in 6 persons with HIV (PWH), where a single infusion of haploidentical related donor NK cells was given plus either IL-2 or N-803 (an IL-15 superagonist). RESULTS: The approach was well tolerated with no unexpected adverse events. We did not pretreat recipients with cyclophosphamide or fludarabine to "make immunologic space," reasoning that PWH on stable antiretroviral treatment remain T-cell depleted in lymphatic tissues. We found donor cells remained detectable in blood for up to 8 days (similar to what is seen in cancer pretreatment with lymphodepleting chemotherapy) and in the lymph nodes and rectum up to 28 days. There was a moderate decrease in the frequency of viral RNA-positive cells in lymph nodes. CONCLUSIONS: There was a moderate decrease in HIV-producing cells in lymph nodes. Further studies are warranted to determine the impact of healthy NK cells on HIV reservoirs and if restoring NK-cell function could be part of an HIV cure strategy. Clinical Trials Registration. NCT03346499 and NCT03899480.

Dual antigen-targeted off-the-shelf NK cells show durable response and prevent antigen escape in lymphoma and leukemia.

Substantial numbers of B cell leukemia and lymphoma patients relapse due to antigen loss or heterogeneity after anti-CD19 chimeric antigen receptor (CAR) T cell therapy. To overcome antigen escape and address antigen heterogeneity, we engineered induced pluripotent stem cell-derived NK cells to express both an NK cell-optimized anti-CD19 CAR for direct targeting and a high affinity, non-cleavable CD16 to augment antibody-dependent cellular cytotoxicity. In addition, we introduced a membrane-bound IL-15/IL-15R fusion protein to promote in vivo persistence. These engineered cells, termed iDuo NK cells, displayed robust CAR-mediated cytotoxic activity that could be further enhanced with therapeutic antibodies targeting B cell malignancies. In multiple in vitro and xenogeneic adoptive transfer models, iDuo NK cells exhibited robust anti-lymphoma activity. Furthermore, iDuo NK cells effectively eliminated both CD19+ and CD19- lymphoma cells and displayed a unique propensity for targeting malignant cells over healthy cells that expressed CD19, features not achievable with anti-CAR19 T cells. iDuo NK cells combined with therapeutic antibodies represent a promising approach to prevent relapse due to antigen loss and tumor heterogeneity in patients with B cell malignancies.

Controlled Stiffness of Direct-Write, Near-Field Electrospun Gelatin Fibers Generates Differences in Tenocyte Morphology and Gene Expression.

Tendinopathy is a leading cause of mobility issues. Currently, the cell-matrix interactions involved in the development of tendinopathy are not fully understood. In vitro tendon models provide a unique tool for addressing this knowledge gap as they permit fine control over biochemical, micromechanical, and structural aspects of the local environment to explore cell-matrix interactions. In this study, direct-write, near-field electrospinning of gelatin solution was implemented to fabricate micron-scale fibrous scaffolds that mimic native collagen fiber size and orientation. The stiffness of these fibrous scaffolds was found to be controllable between 1 MPa and 8 MPa using different crosslinking methods (EDC, DHT, DHT+EDC) or through altering the duration of crosslinking with EDC (1 h to 24 h). EDC crosslinking provided the greatest fiber stability, surviving up to 3 weeks in vitro. Differences in stiffness resulted in phenotypic changes for equine tenocytes with low stiffness fibers (∼1 MPa) promoting an elongated nuclear aspect ratio while those on high stiffness fibers (∼8 MPa) were rounded. High stiffness fibers resulted in the upregulation of matrix metalloproteinase (MMPs) and proteoglycans (possible indicators for tendinopathy) relative to low stiffness fibers. These results demonstrate the feasibility of direct-written gelatin scaffolds as tendon in vitro models and provide evidence that matrix mechanical properties may be crucial factors in cell-matrix interactions during tendinopathy formation.

Spontaneous Spiking Is Governed by Broadband Fluctuations.

Populations of cortical neurons generate rhythmic fluctuations in their ongoing spontaneous activity. These fluctuations can be seen in the local field potential (LFP), which reflects summed return currents from synaptic activity in the local population near a recording electrode. The LFP is spectrally broad, and many researchers view this breadth as containing many narrowband oscillatory components that may have distinct functional roles. This view is supported by the observation that the phase of narrowband oscillations is often correlated with cortical excitability and can relate to the timing of spiking activity and the fidelity of sensory evoked responses. Accordingly, researchers commonly tune in to these channels by narrowband filtering the LFP. Alternatively, neural activity may be fundamentally broadband and composed of transient, nonstationary rhythms that are difficult to approximate as oscillations. In this view, the instantaneous state of the broad ensemble relates directly to the excitability of the local population with no particular allegiance to any frequency band. To test between these alternatives, we asked whether the spiking activity of neocortical neurons in marmoset of either sex is better aligned with the phase of the LFP within narrow frequency bands or with a broadband measure. We find that the phase of broadband LFP fluctuations provides a better predictor of spike timing than the phase after filtering in narrow bands. These results challenge the view of the neocortex as a system composed of narrowband oscillators and supports a view in which neural activity fluctuations are intrinsically broadband.SIGNIFICANCE STATEMENT Research into the dynamical state of neural populations often attributes unique significance to the state of narrowband oscillatory components. However, rhythmic fluctuations in cortical activity are nonstationary and broad spectrum. We find that the timing of spontaneous spiking activity is better captured by the state of broadband fluctuations over any latent oscillatory component. These results suggest narrowband interpretations of rhythmic population activity may be limited, and broader representations may provide higher fidelity in describing moment-to-moment fluctuations in cortical activity.

Exploring cation distribution in ion-exchanged Al,Ga-containing metal-organic frameworks using 17O NMR spectroscopy.

A mixed-metal metal-organic framework, (Al,Ga)-MIL-53, synthesised by post-synthetic ion exchange has been investigated using solid-state nuclear magnetic resonance (NMR) spectroscopy, microscopy and energy dispersive X-ray (EDX) spectroscopy. 17O enrichment during the ion-exchange process enables site specific information on the metal distribution to be obtained. Within this work two ion-exchange processes have been explored. In the first approach (exchange of metals in the framework with dissolved metal salts), 17O NMR spectroscopy reveals the formation of crystallites with a core-shell structure, where the cation exchange takes place on the surface of these materials forming a shell with a roughly equal ratio of Al3+ and Ga3+. For the second approach (exchange of metals between two frameworks), no core-shell structure is observed, and instead crystallites containing a majority of Al3+ are obtained with lower levels of Ga3+. Noticeably, these particles show little variation in the metal cation distribution between crystallites, a result not previously observed for bulk (Al,Ga)-MIL-53 materials. In all cases where ion exchange has taken place NMR spectroscopy reveals a slight preference for clustering of like cations.

Computational NMR investigation of mixed-metal (Al,Sc)-MIL-53 and its phase transitions.

Compositionally complex metal-organic frameworks (MOFs) have properties that depend on local structure that is often difficult to characterise. In this paper a density functional theory (DFT) computational study of mixed-metal (Al,Sc)-MIL-53, a flexible MOF with several different forms, was used to calculate the relative energetics of these forms and to predict NMR parameters that can be used to evaluate whether solid-state NMR spectroscopy can be used to differentiate, identify and characterise the forms adopted by mixed-metal MOFs of different composition. The NMR parameters can also be correlated with structural features in the different forms, giving fundamental insight into the nature and origin of the interactions that affect nuclear spins. Given the complexity of advanced NMR experiments required, and the potential need for expensive and difficult isotopic enrichment, the computational work is invaluable in predicting which experiments and approaches are likely to give the most information on the disorder, local structure and pore forms of these mixed-metal MOFs.

The double-drift illusion biases the marmoset oculomotor system.

The double-drift illusion has two unique characteristics: The error between the perceived and physical position of the stimulus grows over time, and saccades to the moving target land much closer to the physical than the perceived location. These results suggest that the perceptual and saccade targeting systems integrate visual information over different time scales. Functional imaging studies in humans have revealed several potential cortical areas of interest, including the prefrontal cortex. However, we currently lack an animal model to study the neural mechanisms of location perception that underlie the double-drift illusion. To fill this gap, we trained two marmoset monkeys to fixate and then saccade to the double-drift stimulus. In line with human observers for radial double-drift trajectories with fast internal motion, we find that saccade endpoints show a significant bias that is, nevertheless, smaller than the bias seen in human perceptual reports. This bias is modulated by changes in the external and internal speeds of the stimulus. These results demonstrate that the saccade targeting system of the marmoset monkey is influenced by the double-drift illusion.

Conformer-Specific Spectroscopy and IR-Induced Isomerization of a Model γ-Peptide: Ac-γ4-Phe-NHMe.

Single-conformation IR and UV spectroscopy of the prototypical capped γ-peptide Ac-γ4-Phe-NHMe (γ4F) was carried out under jet-cooled conditions in the gas phase in order to understand its innate conformational preferences in the absence of a solvent. We obtained conformer-specific IR and UV spectra and compared the results with calculations to make assignments and explore the differences between the γ2- and γ4-substituted molecules. We found four conformers of γ4F in our experiment. Three conformers form nine-membered hydrogen-bonded rings (C9) enclosed by an NH···O═C H-bond but differing in their phenyl ring positions (a, g+, and g-). The fourth conformer forms a strained seven-membered hydrogen-bonded ring in which the amide groups lie in a nominally anti-parallel arrangement stacked on top of one another (labeled S7). This conformer is a close analogue of the amide-stacked conformer (S) found previously in γ2F, in which the Phe side chain is substituted at the γ2 position, Ac-γ2-Phe-NHMe (J. Am. Chem. Soc. 2009, 131, 14243-14245). IR population transfer spectroscopy was used to determine the fractional abundances of the γ4F conformers in the expansion. A combination of force field and density functional theory calculations is used to map out the conformational potential energy surfaces for γ4F and compare it with its γ2F counterpart. Based on this analysis, the phenyl ring prefers to take up structures that facilitate NH···π interactions in γ4F or avoid phenyl interactions with the C═O group in γ2F. The disconnectivity graph for γ4F reveals separate basins associated with the C9 and amide-stacked conformational families, which are separated by a barrier of about 42 kJ/mol. The overall shape of the potential energy surface bears a resemblance to peptides and proteins that have a misfolding pathway that competes with the formation of the native structure.

Anti-NKG2C/IL-15/anti-CD33 killer engager directs primary and iPSC-derived NKG2C+ NK cells to target myeloid leukemia.

Natural killer (NK) cells mediate the cytolysis of transformed cells and are currently used as an adoptive cellular therapy to treat cancer. Infection with human cytomegalovirus has been shown to expand a subset of "adaptive" NK cells expressing the activation receptor NKG2C that have preferred functional attributes distinct from conventional NK cells. Because NKG2C delivers a strong activating signal to NK cells, we hypothesized that NKG2C could specifically trigger NK-cell-mediated antitumor responses. To elicit a tumor-directed response from NKG2C+ NK cells, we created an anti-NKG2C/IL-15/anti-CD33 killer engager called NKG2C-KE that directs NKG2C+ cells to target CD33+ cells and tumor-associated antigen expressed by acute myelogenous leukemia cells. The NKG2C-KE induced specific degranulation, interferon-γ production, and proliferation of NKG2C-expressing NK cells from patients who reactivated cytomegalovirus after allogeneic transplantation. The NKG2C-KE was also tested in a more homogeneous system using induced pluripotent stem cell (iPSC)-derived NK (iNK) cells that have been engineered to express NKG2C at high levels. The NKG2C-KE triggered iNK-cell-mediated cytotoxicity against CD33+ cells and primary AML blasts. The NKG2C-KE-specific interaction with adaptive NK and NKG2C+ iNK cells represents a new immunotherapeutic paradigm that uniquely engages highly active NK cells to induce cytotoxicity against AML through redirected targeting.

The Paradox of Time in Dynamic Causal Systems.

Recent work has shown that people use temporal information including order, delay, and variability to infer causality between events. In this study, we build on this work by investigating the role of time in dynamic systems, where causes take continuous values and also continually influence their effects. Recent studies of learning in these systems explored short interactions in a setting with rapidly evolving dynamics and modeled people as relying on simpler, resource-limited strategies to grapple with the stream of information. A natural question that arises from such an account is whether interacting with systems that unfold more slowly might reduce the systematic errors that result from these strategies. Paradoxically, we find that slowing the task indeed reduced the frequency of one type of error, albeit at the cost of increasing the overall error rate. To explain these results we posit that human learners analyze continuous dynamics into discrete events and use the observed relationships between events to draw conclusions about causal structure. We formalize this intuition in terms of a novel Causal Event Abstraction model and show that this model indeed captures the observed pattern of errors. We comment on the implications these results have for causal cognition.

Natural killer cells unleashed: Checkpoint receptor blockade and BiKE/TriKE utilization in NK-mediated anti-tumor immunotherapy.

Natural killer (NK) cells have long been known to mediate anti-tumor responses without prior sensitization or recognition of specific tumor antigens. However, the tumor microenvironment can suppress NK cell function resulting in tumor escape and disease progression. Despite recent advances in cytokine therapy and NK cell adoptive transfer, tumor expression of ligands to NK - expressed checkpoint receptors can still suppress NK mediated tumor lysis. This review will explore many of the checkpoint receptors tumors utilize to manipulate the NK cell response as well as some of the current and upcoming pharmacological solutions to limit tumor suppression of NK cell function. Furthermore, we will discuss the potential to use these drugs in combinational therapies with novel antibody reagents such as bi- and tri-specific killer engagers (BiKEs and TriKEs) against tumor-specific antigens to enhance NK cell-mediated tumor rejection.

Following the unusual breathing behaviour of 17O-enriched mixed-metal (Al,Ga)-MIL-53 using NMR crystallography.

The breathing behaviour of 17O-enriched (Al,Ga)-MIL-53, a terephthalate-based metal-organic framework, has been investigated using a combination of solid-state nuclear magnetic resonance (NMR) spectroscopy, powder X-ray diffraction (PXRD) and first-principles calculations. These reveal that the behaviour observed for as-made, calcined, hydrated and subsequently dehydrated mixed-metal MIL-53 materials differs with composition, but cannot be described as the compositionally weighted average of the breathing behaviour seen for the two end members. Although the form of MIL-53 adopted by the as-made material is independent of metal composition, upon calcination, materials with higher levels of Al adopt an open pore (OP) form, as found for the Al end member, but substitution of Ga results in mixed pore materials, with OP and narrow pore (NP) forms co-existing. Although the Ga end member is prone to decomposition under the calcination conditions used, a low level of Al in the starting synthesis (5%) leads to an OP mixed-metal MOF that is stable to calcination. Upon hydration, all materials almost exclusively adopt a closed pore (CP) structure, with strong hydrogen bonding interactions with water leading to two distinct resonances from the carboxylate oxygens in 17O NMR spectra. When dehydrated, different framework structures are found for the two end members, OP for Al-MIL-53 and NP for Ga-MIL-53, with the proportion of NP MOF seen to increase systematically with the Ga content in mixed-metal materials, in contrast to the forms seen upon initial calcination. 17O NMR spectra of mixed-metal MIL-53 materials show an increased preference for clustering of like cations as the Ga content increases. This is not a result of the small-scale dry gel conversion reactions used for enrichment, as a similar cation distribution and clustering is also observed for (Al0.5,Ga0.5)-MIL-53 synthesised hydrothermally and enriched with 17O via post-synthetic steaming.

Nanomechanical Plasmon Spectroscopy of Single Gold Nanoparticles.

We experimentally demonstrate the effect of the localized surface plasmon resonance (LSPR) of a single gold nanoparticle (AuNP) of 100 nm in diameter on the mechanical resonance frequency of a free-standing silicon nitride membrane by means of optomechanical transduction. We discover that a key effect to explain the coupling in these systems is the extinction cross section enhancement due to the excitation of the LSPR at selected wavelengths. In order to validate this coupling, we have developed a fixed wavelength interferometric readout system with an integrated tunable laser source, which allows us to perform the first experimental demonstration of nanomechanical spectroscopy of deposited AuNPs onto the membrane, discerning in between single particles and dimers by the mechanical frequency shift. We have also introduced three-axis mechanical scanners with nanometer-scale resolution in our experimental setup to selectively study single nanoparticles or small clusters. Whereas the single particles are polarization-insensitive, the gold dimers have a clearly defined polarization angle dependency as expected by theory. Finally, we found an unexpected long-distance (∼200 nm) coupling of the LSPR of separated AuNPs coming out from the guided light by the silicon nitride membrane.

Early Reconstitution of NK and γδ T Cells and Its Implication for the Design of Post-Transplant Immunotherapy.

Relapse is the most frequent cause of treatment failure after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Natural killer (NK) cells and γδ T cells reconstitute early after allo-HSCT, contribute to tumor immunosurveillance via major histocompatibility complex-independent mechanisms and do not induce graft-versus-host disease. Here we performed a quantitative and qualitative analysis of the NK and γδ T cell repertoire in healthy individuals, recipients of HLA-matched sibling or unrelated donor allo-HSCT (MSD/MUD-HSCT) and umbilical cord blood-HSCT (UCB-HSCT). NK cells are present at high frequencies in all allo-HSCT recipients. Immune reconstitution (IR) of vδ2+ cells depended on stem cell source. In MSD/MUD-HSCT recipients, vδ2+ comprise up to 8% of the total lymphocyte pool, whereas vδ2+ T cells are barely detectable in UCB-HSCT recipients. Vδ1+ IR was driven by CMV reactivation and was comparable between MSD/MUD-HSCT and UCB-HSCT. Strategies to augment NK cell mediated tumor responses, similar to IL-15 and antibodies, also induced vδ2+ T cell responses against a variety of different tumor targets. Vδ1+ γδ T cells were induced less by these same stimuli. We also identified elevated expression of the checkpoint inhibitory molecule TIGIT (T cell Ig and ITIM domain), which is also observed on tumor-infiltrating lymphocytes and epidermal γδ T cells. Collectively, these data show multiple strategies that can result in a synergized NK and γδ T cell antitumor response. In the light of recent developments of low-toxicity allo-HSCT platforms, these interventions may contribute to the prevention of early relapse.

A Conserved HIV-1-Derived Peptide Presented by HLA-E Renders Infected T-cells Highly Susceptible to Attack by NKG2A/CD94-Bearing Natural Killer Cells.

Major histocompatibility class I (MHC-I)-specific inhibitory receptors on natural killer (NK) cells (iNKRs) tolerize mature NK cell responses toward normal cells. NK cells generate cytolytic responses to virus-infected or malignant target cells with altered or decreased MHC-I surface expression due to the loss of tolerizing ligands. The NKG2A/CD94 iNKR suppresses NK cell responses through recognition of the non-classical MHC-I, HLA-E. We used HIV-infected primary T-cells as targets in an in vitro cytolytic assay with autologous NK cells from healthy donors. In these experiments, primary NKG2A/CD94(+) NK cells surprisingly generated the most efficient responses toward HIV-infected T-cells, despite high HLA-E expression on the infected targets. Since certain MHC-I-presented peptides can alter recognition by iNKRs, we hypothesized that HIV-1-derived peptides presented by HLA-E on infected cells may block engagement with NKG2A/CD94, thereby engendering susceptibility to NKG2A/CD94(+) NK cells. We demonstrate that HLA-E is capable of presenting a highly conserved peptide from HIV-1 capsid (AISPRTLNA) that is not recognized by NKG2A/CD94. We further confirmed that HLA-C expressed on HIV-infected cells restricts attack by KIR2DL(+) CD56(dim) NK cells, in contrast to the efficient responses by CD56(bright) NK cells, which express predominantly NKG2A/CD94 and lack KIR2DLs. These findings are important since the use of NK cells was recently proposed to treat latently HIV-1-infected patients in combination with latency reversing agents. Our results provide a mechanistic basis to guide these future clinical studies, suggesting that ex vivo-expanded NKG2A/CD94(+) KIR2DL(-) NK cells may be uniquely beneficial.

Increasing Spontaneous Retinal Activity before Eye Opening Accelerates the Development of Geniculate Receptive Fields.

Visually evoked activity is necessary for the normal development of the visual system. However, little is known about the capacity for patterned spontaneous activity to drive the maturation of receptive fields before visual experience. Retinal waves provide instructive retinotopic information for the anatomical organization of the visual thalamus. To determine whether retinal waves also drive the maturation of functional responses, we increased the frequency of retinal waves pharmacologically in the ferret (Mustela putorius furo) during a period of retinogeniculate development before eye opening. The development of geniculate receptive fields after receiving these increased neural activities was measured using single-unit electrophysiology. We found that increased retinal waves accelerate the developmental reduction of geniculate receptive field sizes. This reduction is due to a decrease in receptive field center size rather than an increase in inhibitory surround strength. This work reveals an instructive role for patterned spontaneous activity in guiding the functional development of neural circuits.

Adaptive Natural Killer Cell and Killer Cell Immunoglobulin-Like Receptor-Expressing T Cell Responses are Induced by Cytomegalovirus and Are Associated with Protection against Cytomegalovirus Reactivation after Allogeneic Donor Hematopoietic Cell Transplantation.

Cytomegalovirus (CMV) reactivates in >30% of CMV-seropositive patients after allogeneic hematopoietic cell transplantation (HCT). Previously, we reported an increase of natural killer (NK) cells expressing NKG2C, CD57, and inhibitory killer cell immunoglobulin-like receptors (KIRs) in response to CMV reactivation after HCT. These NK cells persist after the resolution of infection and display "adaptive" or memory properties. Despite these findings, the differential impact of persistent/inactive versus reactivated CMV on NK versus T cell maturation after HCT from different graft sources has not been defined. We compared the phenotype of NK and T cells from 292 recipients of allogeneic sibling (n = 118) or umbilical cord blood (UCB; n = 174) grafts based on recipient pretransplantation CMV serostatus and post-HCT CMV reactivation. This cohort was utilized to evaluate CMV-dependent increases in KIR-expressing NK cells exhibiting an adaptive phenotype (NKG2C(+)CD57(+)). Compared with CMV-seronegative recipients, those who reactivated CMV had the highest adaptive cell frequencies, whereas intermediate frequencies were observed in CMV-seropositive recipients harboring persistent/nonreplicating CMV. The same effect was observed in T cells and CD56(+) T cells. These adaptive lymphocyte subsets were increased in CMV-seropositive recipients of sibling but not UCB grafts and were correlated with lower rates of CMV reactivation (sibling 33% versus UCB 51%; P < .01). These data suggest that persistent/nonreplicating recipient CMV induces rapid production of adaptive NK and T cells from mature cells from sibling but not UCB grafts. These adaptive lymphocytes are associated with protection from CMV reactivation.

Dynamic hydrogen bonding and DNA flexibility in minor groove binders: molecular dynamics simulation of the polyamide f-ImPyIm bound to the Mlu1 (MCB) sequence 5'-ACGCGT-3' in 2:1 motif.

Molecular dynamics simulations of the DNA 10-mer 5'-CCACGCGTGG-3' alone and complexed with the formamido-imidazole-pyrrole-imidazole (f-ImPyIm) polyamide minor groove binder in a 2:1 fashion were conducted for 50 ns using the pbsc0 parameters within the AMBER 12 software package. The change in DNA structure upon binding of f-ImPyIm was evaluated via minor groove width and depth, base pair parameters of Slide, Twist, Roll, Stretch, Stagger, Opening, Propeller, and x-displacement, dihedral angle distributions of ζ, ε, α, and γ determined using the Curves+ software program, and hydrogen bond formation. The dynamic hydrogen bonding between the f-ImPyIm and its cognate DNA sequence was compared to the static image used to predict sequence recognition by polyamide minor groove binders. Many of the predicted hydrogen bonds were present in less than 50% of the simulation; however, persistent hydrogen bonds between G5/15 and the formamido group of f-ImPyIm were observed. It was determined that the DNA is wider in the Complex than without the polyamide binder; however, there is flexibility in this particular sequence, even in the presence of the f-ImPyIm as evidenced by the range of minor groove widths the DNA exhibits and the dynamics of the hydrogen bonding that binds the two f-ImPyIm ions to the minor groove. The Complex consisting of the DNA and the 2 f-ImPyIm binders shows slight fraying of the 5' end of the 10-mer at the end of the simulation, but the portion of the oligomer responsible for recognition and binding is stable throughout the simulation. Several structural changes in the Complex indicate that minor groove binders may have a more active role in inhibiting transcription than just preventing binding of important transcription factors.

Instrumental variable estimation for functional concurrent regression models.

In this work we propose a functional concurrent regression model to estimate labor supply elasticities over the years 1988 through 2014 using Current Population Survey data. Assuming, as is common, that individuals' wages are endogenous, we introduce instrumental variables in a two-stage least squares approach to estimate the desired labor supply elasticities. Furthermore, we tailor our estimation method to sparse functional data. Though recent work has incorporated instrumental variables into other functional regression models, to our knowledge this has not yet been done in the functional concurrent regression model, and most existing literature is not suited for sparse functional data. We show through simulations that this two-stage least squares approach greatly eliminates the bias introduced by a naive model (i.e. one that does not acknowledge endogeneity) and produces accurate coefficient estimates for moderate sample sizes.