Structural and functional definition of a vulnerable site on the hemagglutinin of highly pathogenic avian influenza A virus H5N1.

Most neutralizing antibodies against highly pathogenic avian influenza A virus H5N1 recognize the receptor-binding site (RBS) on the globular head domain and the stem of H5N1 hemagglutinin (HA). Through comprehensive analysis of multiple human protective antibodies, we previously identified four vulnerable sites (VS1-VS4) on the globular head domain. Among them, the VS1, occupying the opposite side of the RBS on the same HA, was defined by the epitope of antibody 65C6. In this study, we report the crystal structures of two additional human H5N1 antibodies isolated from H5N1-infected individuals, 3C11 and AVFluIgG01, bound to the head at 2.33- and 2.30-Å resolution, respectively. These two new antibody epitopes have large overlap with and extend beyond the original VS1. Site-directed mutagenesis experiments identified eight pivotal residues (Ser-126b, Lys-165, Arg-166, Ser-167, Tyr-168, Asn-169, Thr-171, and Asn-172) critical for 65C6-, 3C11-, and AVFluIgG01-binding and neutralization activities. These residues formed a unique "Y"-shaped surface on H5N1 globular head and are highly conserved among H5N1 viruses. Our results further support the existence of a vulnerable site distinct from the RBS and the stem region of H5N1 HA, and future design of immunogens should take this particular site into consideration.

Complementary recognition of the receptor-binding site of highly pathogenic H5N1 influenza viruses by two human neutralizing antibodies.

The highly pathogenic avian influenza virus H5N1 is a major threat to global public health and therefore a high-priority target of current vaccine development. The receptor-binding site (RBS) on the globular head of hemagglutinin (HA) in the viral envelope is one of the major target sites for antibody recognition against H5N1 and other influenza viruses. Here, we report the identification and characterization of a pair of human RBS-specific antibodies, designated FLD21.140 and AVFluIgG03, that are mutually complementary in their neutralizing activities against a diverse panel of H5N1 viruses. Crystallographic analysis and site-directed mutagenesis revealed that the two antibodies share a similar RBS-binding mode, and their individual specificities are governed by residues at positions 133a, 144, and 145. Specifically, FLD21.140 preferred Leu-133a/Lys-144/Ser-145, whereas AVFluIgG03 favored Ser-133a/Thr-144/Pro-145 residue triplets, both of which perfectly matched the most prevalent residues in viruses from epidemic-originating regions. Of note, according to an analysis of 3758 H5 HA sequences available in the Influenza Virus Database at the National Center for Biotechnology Information, the residues Leu-133a/Ser-133a and Ser-145/Pro-145 constituted more than 87.6 and 99.3% of all residues at these two positions, respectively. Taken together, our results provide a structural understanding for the neutralizing complementarity of these two antibodies and improve our understanding of the RBS-specific antibody response against H5N1 infection in humans.

Double dissociation of configural and featural face processing on P1 and P2 components as a function of spatial attention.

Face recognition relies on both configural and featural processing. Previous research has shown that P1 is sensitive to configural face processing, but it is unclear whether any component is sensitive to featural face processing; moreover, if there is such a component, its temporal sequence relative to P1 is unknown. Thus, to avoid confounding physical stimuli differences between configural and featural face processing on ERP components, a spatial attention paradigm was employed by instructing participants to attend an image stream (faces and houses) or an alphanumeric character stream. The interaction between attention and face processing type on P1 and P2 components indicates different mechanisms of configural and featural face processing as a function of spatial attention. The steady-state visual evoked potential (SSVEP) results clearly demonstrated that participants could selectively attend to different streams of information. Importantly, configural face processing elicited a larger posterior P1 (approximately 128 ms) than featural face processing, whereas P2 (approximately 248 ms) was greater for featural than for configural face processing under attended condition. The interaction between attention and face processing type (configural vs. featural) on P1 and P2 components indicates that there are different mechanisms of configural and featural face processing operating as functions of spatial attention. While the P1 result confirms previous findings separating configural and featural face processing, the newly observed P2 finding in the present study extends this separation to a double dissociation. Therefore, configural and featural face processing are modulated differently by spatial attention, and configural face processing precedes featural face processing.

Long-Term Exposure to High Altitude Affects Conflict Control in the Conflict-Resolving Stage.

The neurocognitive basis of the effect of long-term high altitude exposure on conflict control is unclear. Event related potentials (ERPs) were recorded in a flanker task to investigate the influence of high altitude on conflict control in the high-altitude group (who had lived at high altitude for three years but were born at low altitude) and the low-altitude group (living in low altitude only). Although altitude effect was not significant at the behavioral level, ERPs showed cognitive conflict modulation. The interaction between group and trial type was significant: P3 amplitude was greater in the low-altitude group than in the high-altitude group in the incongruent trial. This result suggests that long-term exposure to high altitude affects conflict control in the conflict-resolving stage, and that attentional resources are decreased to resist the conflict control in the high-altitude group.

BOLD signal change and contrast reversing frequency: an event-related fMRI study in human primary visual cortex.

It is believed that human primary visual cortex (V1) increases activity with increasing temporal frequency of a visual stimulus. Two kinds of visual stimulus were used in the previous studies, one is patterned-flash stimulus with a fixed onset period and an increasing average luminance with the increase of temporal frequency, the other is contrast reversing flickering checkerboard or grating with a constant average luminance across different temporal frequencies. That hemodynamic responses change as a function of reversal frequency of contrast reversing checkerboard is at odds with neurophysiological studies in animals and neuroimaging studies in humans. In the present study, we addressed the relationship between reversal frequency of contrast reversing checkerboard and hemodynamic response in human V1 using an event-related experimental paradigm and found that the transient characteristics of blood oxygenation level dependent response in human V1 depended very little on the reversal frequency of a contrast reversing checkerboard.

Neural adaptation provides evidence for categorical differences in processing of faces and Chinese characters: an ERP study of the N170.

Whether face perception involves domain-specific or domain-general processing is an extensively debated issue. Relative to non-face objects and alphabetical scripts, Chinese characters provide a good contrast to faces because of their structural configuration, requirement for high level of visual expertise to literate Chinese people, and unique appearance and identity for each individual stimulus. To examine potential categorical differences in their neural processing, event-related potentials (ERPs) were recorded to blocked face and Chinese character stimuli. Fast adaptation method was applied to better control for the low-level stimulus difference between faces and Chinese characters. Participants were required to respond to the color of the outer frame in which these stimuli were presented, at either a fast (ISI 650 ms) or slow (ISI 1300 ms) rate, and with an orientation that was either the same or alternated between upright and inverted. Faces elicited a larger and later N170 relative to characters, but the N170 was more left-lateralized for characters relative to the faces. Adaptation-by-rate and adaptation-by-orientation effects were observed on the amplitude of N170, and both were more pronounced for faces relative to characters. Inverted stimuli elicited a later N170 relative to upright stimuli, without amplitude change, and this inversion effect was more pronounced for faces relative to characters. Moreover, faces elicited a larger and later P1 and a larger adaptation-by-rate effect on P1 relative to characters. The adaptation-by-orientation effect was illustrated by a larger P1 under the same relative to the alternated orientation condition. Therefore, evidence from the amplitude and the lateralization of N170, the stimulus inversion effect on N170 latency, and the neural adaptation between faces and Chinese characters on P1 and N170 components support the notion that the processing of faces and Chinese characters involve categorically different neural mechanisms.