Biology subjects

White, A. L.

Publications and source records attributed to White, A. L..

8 recordsLinked to original sources

bioRxiv2025

A buffer-tuning strategy to profile domain-specific activity of chimeric I-TevI/CRISPR gene editors in vitro

Protein-DNA interactions can be manipulated in vitro by changing buffer conditions. Here, we develop a methodology to map the cleavage preferences of chimeric gene editors that are fusions of the I-TevI nuclease domain to CRIPSR nucleases by manipulating in vitro salt concentrations. We found that DNA cleavage by the I-TevI (Tev) nuclease domain at CNNNG sites was de-coupled from the gRNA-targeted site in low salt buffers. For TevCas12a, this non-targeted cleavage activity was enriched at Tev CNNNG cleavage motifs optimally positioned within a 30-bp window upstream of a Cas12a TTTV PAM site. Non-targeted cleavage did not require Cas12a nuclease activity or specific Cas12a gRNA targeting. Similar non-targeted products were observed in low salt buffer conditions for TevSaCas9, Tev-meganuclease and Tev-zinc finger editors. Cas12a and SaCas9 activity at gRNA-directed sites and sites with multiple mismatches were also sensitive to buffer salt concentration. Oxford Nanopore sequencing revealed a remarkably similar Tev CNNNG cleavage preference at different salt concentrations and in different fusion contexts, emphasizing the robustness and specificity of Tev activity. More generally, our work highlights the sensitivity of gene editors to in vitro reaction conditions and how these conditions can be leveraged to functionally dissect the activity of individual domains of chimeric gene editors.

biochemistry↗

bioRxiv2025

Simultaneous cortical responses to multiple written words

We have learned much about the brain regions that support reading by measuring neuronal responses to single words, but we know little about how the brain processes multiple words simultaneously. This fMRI study fills that gap by varying the number of English words presented while holding the amount of visual stimulation constant. We adapted the "simultaneous suppression" paradigm, which has demonstrated that the response to multiple stimuli presented simultaneously is typically smaller than the sum of responses to the same stimuli presented sequentially. On each trial, participants viewed a rapid sequence of three frames. Each frame contained two character strings, most of which were pseudo-letters with visual features matched to familiar letters. The experimental conditions differed in the number of words in the sequence: zero words; one word; two words sequentially; or two words simultaneously. Behavioral task accuracy was worse for detecting two words presented simultaneously than sequentially. BOLD responses increased linearly with the number of words presented in several reading-related regions of the left hemisphere: text-selective occipito-temporal regions, the superior temporal sulcus, the intraparietal sulcus, and the inferior frontal sulcus. In all of those regions, responses did not differ between sequential and simultaneous presentation of two words. Nonetheless, the sensitivity of ventral temporal text-selective regions to the lexical frequencies of two words was attenuated by simultaneous presentation. To account for these patterns of activity and task performance, we suggest that the reading network can detect two strings of letters simultaneously, but there is interference during lexical access.

neuroscience↗

bioRxiv2025

Transcriptomic Analysis of CAD Cell Differentiation

CAD cells were derived from Cath.a cells, a mouse central nervous system catecholaminergic cell line. Serum-starved CAD cells undergo morphological changes and resemble isolated neurons when observed by microscopy. We carried out an RNAseq transcriptomic analysis to examine differentiated CAD cells for expression signatures related to neuronal functions, identifying [~]1900 transcripts whose expression changed with differentiation. Pathview analysis identified [~]80 KEGG pathway gene sets that were differentially expressed, including upregulation of at least 13 neuron-related pathways. This dataset can be explored more deeply, allowing further investigation into expression changes relevant to studying neuronal functions in this easy-to-culture model system.

neuroscience↗

bioRxiv2023

Voluntary effort to read reshapes stimulus selectivity in the visual word form area and increases its communication with the language network

Reading depends on a brain region known as the "visual word form area" (VWFA) in left ventral occipito-temporal cortex. This regions function is debated because its stimulus selectivity is not absolute, it is modulated by a variety of task demands, and it is inconsistently localized. We used fMRI to characterize the combination of sensory and cognitive factors that activate word-responsive regions that we precisely localized in 16 adult humans (4 male). We then presented three types of character strings: English words, pseudowords, and unfamiliar characters with matched visual features. Participants performed three different tasks while viewing those stimuli: detecting real words, detecting color in the characters, and detecting color in the fixation mark. There were three primary findings about the VWFAs response: (1) It preferred letter strings over unfamiliar characters even when the stimuli were ignored during the fixation task; (2) Compared to those baseline responses, engaging in the word reading task enhanced the response to words but suppressed the response to unfamiliar characters. (3) Attending to the stimuli to judge their font color had little effect on the response magnitudes. Thus, the VWFA is uniquely modulated by a cognitive signal that is specific to voluntary linguistic processing and is not additive. Functional connectivity analyses revealed that communication between the VWFA and a left frontal language area increased when the participant engaged in the linguistic task. We conclude that the VWFA is inherently selective for familiar orthography, but it falls under control of the language network when the task demands it. Significance StatementThe function of the "visual word form area" (VWFA) is controversial. Some researchers emphasize its bottom-up visual selectivity for words, hence the regions common name. Others argue that its activity is explained by feedback from regions that control attention or language. To seek clarity, we investigated what drives the VWFA: seeing words, attending visually to words, or trying to read words. None of those factors was sufficient on its own. Our results support a hybrid model: the VWFA has inherent selectivity for words, but its function is reshaped by voluntary language processing. Thus, with an integrated analysis of sensory inputs, task demands, and network connectivity, we provide some resolution to debates about this important region.

neuroscience↗

bioRxiv2022

Engaging in word recognition elicits highly specific modulations in visual cortex

A persons cognitive state determines how their brain responds to visual stimuli. The most common such effect is a response enhancement when stimuli are task-relevant and attended rather than ignored. In this fMRI study, we report a surprising twist on such attention effects in the visual word form area (VWFA), a region that plays a key role in reading. We presented participants with strings of letters and visually similar shapes which were either relevant for a specific task (lexical decision or gap localization) or ignored (during a fixation dot color task). In the VWFA, the enhancement of responses to attended stimuli occurred only for letter strings, whereas the shapes evoked smaller responses when attended than when ignored. The enhancement of VWFA activity was accompanied by strengthened functional connectivity with higher-level language regions. These task-dependent modulations of response magnitude and functional connectivity were specific to the VWFA and absent in the rest of visual cortex. We suggest that language regions send targeted excitatory feedback into the VWFA only when the observer is trying to read. This feedback enables the discrimination of familiar and nonsense words, and is distinct from generic effects of visual attention.

neuroscience↗

bioRxiv2021

Oculomotor freezing tracks perception and is immune to decision bias

The appearance of a salient stimulus rapidly inhibits saccadic eye movements. Curiously, this "oculomotor freezing" reflex is triggered only by stimuli that the observer reports seeing. It remains unknown, however, if oculomotor freezing is linked to the observers sensory experience, or their decision that a stimulus was present. To dissociate between these possibilities, we manipulated decision criterion via monetary payoffs and stimulus probability in a detection task. These manipulations greatly shifted observers decision criteria but did not affect the degree to which microsaccades were inhibited by stimulus presence. Moreover, the link between oculomotor freezing and explicit reports of stimulus presence was stronger when the criterion was conservative rather than liberal. We conclude that the sensory threshold for oculomotor freezing is independent of decision bias. Provided that conscious experience is also unaffected by such bias, oculomotor freezing is an implicit indicator of sensory awareness. New & NoteworthySometimes a visual stimulus reaches awareness, and sometimes it does not. To understand why, we need objective, bias-free measures of awareness. We discovered that a reflexive freezing of small eye movements indicates when an observer detects a stimulus. Furthermore, when we biased observers decisions to report seeing the stimulus, the oculomotor reflex was unaltered. This suggests that the threshold for conscious perception is independent of the decision criterion and is revealed by oculomotor freezing.

neuroscience↗

bioRxiv2020

Divided Attention in Perception:A Unified Analysis of Dual-Task Deficits and Congruency Effects

How well can one perceive simultaneous stimuli at two widely spaced visual locations? Are the stimuli processed independently? If not, does the dependency affect perception, disrupt signals in later stages, or both? To address these questions, we measured effects of divided attention using a dual-task paradigm with stimuli presented in noise on either side of fixation. This paradigm was applied to detecting Gabor patches and to the semantic categorization of words. We measured dual-task deficits which are a decline in mean performance for a dual task compared to a single task. There was such a deficit for categorizing two words but relatively little deficit for detecting two Gabors. We also measured congruency effects which are when performance at one location depends on whether the stimulus at the other location requires the same response. There was such a congruency effect for detecting two Gabors but relatively little congruency effect for categorizing two words. Further experiments were consistent with the dual-task deficit in word categorization being perceptual, but the congruency effect in Gabor detection being due to later processes. Results of additional experiments showed that the congruency effect was consistent with either graded selection errors or with all-or-none selection followed by graded interactive processing. To answer our opening question: for Gabor detection, perceptual processes were largely independent but later processes caused congruency effects; for word categorization, perceptual processes had capacity limits but even in combination with later processes caused relatively little congruency effects. In summary, there was evidence for two different kinds of dependency. Such complementary dependencies are inconsistent with theories of divided attention that depend on a single dependency such as a single resource or single source of interactive processing.

neuroscience↗

bioRxiv2018

Parallel spatial channels converge at a bottleneck in anterior word-selective cortex

In most environments, the visual system is confronted with many relevant objects simultaneously. That is especially true during reading. However, behavioral data demonstrate that a serial bottleneck prevents recognition of more than one word at a time. We used functional magnetic resonance imaging (fMRI) to investigate how parallel spatial channels of visual processing converge into a serial bottleneck for word recognition. Observers viewed pairs of words presented simultaneously. We found that retinotopic cortex processed the two words in parallel spatial channels, one in each contralateral hemisphere. Responses were higher for attended words than ignored words, but were not reduced when attention was divided, even though behavioral performance suffered greatly. We then analyzed two word-selective regions along the occipito-temporal sulcus (OTS) of both hemispheres (i.e., sub-regions of visual word form area, VWFA). Unlike retinotopic cortex, each word-selective region responded to words on both sides of fixation. Nonetheless, a single region in the left hemisphere (VWFA-1 in posterior OTS) contained spatial channels for both hemifields that were independently modulated by selective attention. Thus, the left posterior VWFA supports parallel processing of multiple words. In contrast, a more anterior word-selective region in the left hemisphere (VWFA-2 in mid-OTS) showed limited spatial and attentional selectivity, consistent with activity of a single channel. Therefore, the visual system can process two words in parallel up to a late stage in the ventral stream. The transition from two parallel channels to a single channel in more anterior regions is consistent with the observed bottleneck in behavior.

neuroscience↗