Multiciliated ependymal cells: an update on biology and pathology in the adult brain.

Mature multiciliated ependymal cells line the cerebral ventricles where they form a partial barrier between the cerebrospinal fluid (CSF) and brain parenchyma and regulate local CSF microcirculation through coordinated ciliary beating. Although the ependyma is a highly specialized brain interface with barrier, trophic, and perhaps even regenerative capacity, it remains a misfit in the canon of glial neurobiology. We provide an update to seminal reviews in the field by conducting a scoping review of the post-2010 mature multiciliated ependymal cell literature. We delineate how recent findings have either called into question or substantiated classical views of the ependymal cell. Beyond this synthesis, we document the basic methodologies and study characteristics used to describe multiciliated ependymal cells since 1980. Our review serves as a comprehensive resource for future investigations of mature multiciliated ependymal cells.

Naturally occurring splice variants dissect the functional domains of BHC80 and emphasize the need for RNA analysis.

Pathogenic PHF21A variation causes PHF21A-related neurodevelopmental disorders (NDDs). Although amorphic alleles, including haploinsufficiency, have been established as a disease mechanism, increasing evidence suggests that missense variants as well as frameshift variants extending the BHC80 carboxyl terminus also cause disease. Expanding on these, we report a proposita with intellectual disability and overgrowth and a novel de novo heterozygous PHF21A splice variant (NM_001352027.3:c.[153+1G>C];[=]) causing skipping of exon 6, which encodes an in-frame BHC80 deletion (p.(Asn30_Gln51del)). This deletion disrupts a predicted leucine zipper domain and implicates this domain in BHC80 function and as a target of variation causing PHF21A-related NDDs. This extension of understanding emphasizes the application of RNA analysis in precision genomic medicine practice.

NOTCH1 loss of the TAD and PEST domain: An antimorph?

The Notch proteins play key roles in cell fate determination during development. Germline pathogenic variants in NOTCH1 predispose to a spectrum of cardiovascular malformations including Adams-Oliver syndrome and a wide variety of isolated complex and simple congenital heart defects. The intracellular C-terminus of the single-pass transmembrane receptor encoded by NOTCH1 contains a transcriptional activating domain (TAD) required for target gene activation and a PEST domain (a sequence rich in proline, glutamic acid, serine, and threonine), regulating protein stability and turnover. We present a patient with a novel variant encoding a truncated NOTCH1 protein without the TAD and PEST domain (NM_017617.4: c.[6626_6629del];[=], p.(Tyr2209CysfsTer38)) and extensive cardiovascular abnormalities consistent with a NOTCH1-mediated mechanism. This variant fails to promote transcription of target genes as assessed by luciferase reporter assay. Given the roles of the TAD and PEST domains in NOTCH1 function and regulation, we hypothesize that loss of both the TAD and the PEST domain results in a stable, loss-of-function protein that acts as an antimorph through competition with wild-type NOTCH1.

The Clinical Variant Analysis Tool: Analyzing the evidence supporting reported genomic variation in clinical practice.

PURPOSE: Genomic test results, regardless of laboratory variant classification, require clinical practitioners to judge the applicability of a variant for medical decisions. Teaching and standardizing clinical interpretation of genomic variation calls for a methodology or tool. METHODS: To generate such a tool, we distilled the Clinical Genome Resource framework of causality and the American College of Medical Genetics/Association of Molecular Pathology and Quest Diagnostic Laboratory scoring of variant deleteriousness into the Clinical Variant Analysis Tool (CVAT). Applying this to 289 clinical exome reports, we compared the performance of junior practitioners with that of experienced medical geneticists and assessed the utility of reported variants. RESULTS: CVAT enabled performance comparable to that of experienced medical geneticists. In total, 124 of 289 (42.9%) exome reports and 146 of 382 (38.2%) reported variants supported a diagnosis. Overall, 10.5% (1 pathogenic [P] or likely pathogenic [LP] variant and 39 variants of uncertain significance [VUS]) of variants were reported in genes without established disease association; 20.2% (23 P/LP and 54 VUS) were in genes without sufficient phenotypic concordance; 7.3% (15 P/LP and 13 VUS) conflicted with the known molecular disease mechanism; and 24% (91 VUS) had insufficient evidence for deleteriousness. CONCLUSION: Implementation of CVAT standardized clinical interpretation of genomic variation and emphasized the need for collaborative and transparent reporting of genomic variation.

Mesenteric cysts, lymphatic leak, and cerebral cavernous malformation in a proband with KRIT1-related disease.

Cerebral cavernous malformations (CCMs) of the central nervous system arise sporadically or secondary to genomic variation. Established genetic etiologies include deleterious variants in KRIT1 (CCM1), malcavernin (CCM2), and PDCD10 (CCM3). KRIT1-related disease has not been described in conjunction with lymphatic defects, although lymphatic defects with abnormal endothelial cell junctions have been observed in mice deficient in HEG1-KRIT1 signaling. We report a proband with CCMs, multiple chylous mesenteric cysts, and chylous ascites with leaky lymphatic vasculature. Clinical short-read exome sequencing detected a disease-associated KRIT1 variant (NM_194456.1:c.[1927C>T];[=], p.(Gln643*)). We postulate an expansion of KRIT1-related disease to include lymphatic malformations and lymphatic endothelial dysfunction.

Long-read genome sequencing resolves a complex 13q structural variant associated with syndromic anophthalmia.

Microphthalmia, anophthalmia, and coloboma (MAC) are a heterogeneous spectrum of anomalous eye development and degeneration with genetic and environmental etiologies. Structural and copy number variants of chromosome 13 have been implicated in MAC; however, the specific loci involved in disease pathogenesis have not been well-defined. Herein we report a newborn with syndromic degenerative anophthalmia and a complex de novo rearrangement of chromosome 13q. Long-read genome sequencing improved the resolution and clinical interpretation of a duplication-triplication/inversion-duplication (DUP-TRP/INV-DUP) and terminal deletion. Sequence features at the breakpoint junctions suggested microhomology-mediated break-induced replication (MMBIR) of the maternal chromosome as the origin. Comparing this rearrangement to previously reported copy number alterations in 13q, we refine a putative dosage-sensitive critical region for MAC that might provide new insights into its molecular etiology.

An infant with congenital respiratory insufficiency and diaphragmatic paralysis: A novel BICD2 phenotype?

Monoallelic pathogenic variants in BICD2 are associated with autosomal dominant Spinal Muscular Atrophy Lower Extremity Predominant 2A and 2B (SMALED2A, SMALED2B). As part of the cellular vesicular transport, complex BICD2 facilitates the flow of constitutive secretory cargoes from the trans-Golgi network, and its dysfunction results in motor neuron loss. The reported phenotypes among patients with SMALED2A and SMALED2B range from a congenital onset disorder of respiratory insufficiency, arthrogryposis, and proximal or distal limb weakness to an adult-onset disorder of limb weakness and contractures. We report an infant with congenital respiratory insufficiency requiring mechanical ventilation, congenital diaphragmatic paralysis, decreased lung volume, and single finger camptodactyly. The infant displayed appropriate antigravity limb movements but had radiological, electrophysiological, and histopathological evidence of myopathy. Exome sequencing and long-read whole-genome sequencing detected a novel de novo BICD2 variant (NM_001003800.1:c.[1543G>A];[=]). This is predicted to encode p.(Glu515Lys); p.Glu515 is located in the coiled-coil 2 mutation hotspot. We hypothesize that this novel phenotype of diaphragmatic paralysis without clear appendicular muscle weakness and contractures of large joints is a presentation of BICD2-related disease.

Impact of aging on inflammatory and immune responses during elastin peptide-induced murine emphysema.

Deterioration of lung functions and degradation of elastin fibers with age are accelerated during chronic obstructive pulmonary disease (COPD). Excessive genesis of soluble elastin peptides (EP) is a key factor in the pathophysiology of COPD. We have previously demonstrated that 6-wk-old mice exhibited emphysematous structural changes associated with proinflammatory immune response after EP instillation. In this study, we investigated the consequences of aging on inflammatory, immune, and histological criteria associated with murine emphysema progression after EP exposure. Young (6 wk old) and elderly (15 mo old) C57BL/6J mice were endotracheally instilled with EP, and, at various time points after treatment, the inflammatory cell profiles from bronchoalveolar lavage fluids (BALF) and the T-lymphocyte phenotypes, at local and systemic levels, were analyzed by flow cytometry. Lungs were also prepared to allow morphological and histological analysis by confocal microscopy. Elderly mice exhibited an earlier development of pulmonary emphysema, characterized by an increase of the inflammatory and lymphocytic infiltrates, extracellular matrix breakdown, and airspace enlargement compared with young mice. This age-dependent parenchymal tissue remodeling was associated with an increase of the matrix metalloproteinase expressions and desmosine levels in BALF and/or sera of EP-treated mice. In addition, both the proportion of CD4+CD28- and CD8+CD28- T cells in the tissues of EP-treated mice and the interferon-γ levels in the EP-specific memory T-cell clones were significantly higher in elderly versus younger mice. This study demonstrates that aging accelerates emphysema development and that this effect is linked to increased EP production and their effects on inflammatory and immune response.

Optical Control of Metal Ion Probes in Cells and Zebrafish Using Highly Selective DNAzymes Conjugated to Upconversion Nanoparticles.

Spatial and temporal distributions of metal ions in vitro and in vivo are crucial in our understanding of the roles of metal ions in biological systems, and yet there is a very limited number of methods to probe metal ions with high space and time resolution, especially in vivo. To overcome this limitation, we report a Zn2+-specific near-infrared (NIR) DNAzyme nanoprobe for real-time metal ion tracking with spatiotemporal control in early embryos and larvae of zebrafish. By conjugating photocaged DNAzymes onto lanthanide-doped upconversion nanoparticles (UCNPs), we have achieved upconversion of a deep tissue penetrating NIR 980 nm light into 365 nm emission. The UV photon then efficiently photodecages a substrate strand containing a nitrobenzyl group at the 2'-OH of adenosine ribonucleotide, allowing enzymatic cleavage by a complementary DNA strand containing a Zn2+-selective DNAzyme. The product containing a visible FAM fluorophore that is initially quenched by BHQ1 and Dabcyl quenchers is released after cleavage, resulting in higher fluorescent signals. The DNAzyme-UCNP probe enables Zn2+ sensing by exciting in the NIR biological imaging window in both living cells and zebrafish embryos and detecting in the visible region. In this study, we introduce a platform that can be used to understand the Zn2+ distribution with spatiotemporal control, thereby giving insights into the dynamical Zn2+ ion distribution in intracellular and in vivo models.

Hormone replacement therapy, mammographic density, and breast cancer risk: a cohort study.

PURPOSE: Hormone replacement therapy (HRT) use increases breast cancer risk and mammographic density (MD). We examine whether MD mediates or modifies the association of HRT with the breast cancer. METHODS: For the 4,501 participants in the Danish diet, cancer and health cohort (1993-1997) who attended mammographic screening in Copenhagen (1993-2001), MD (mixed/dense or fatty) was assessed at the first screening after cohort entry. HRT use was assessed by questionnaire and breast cancer diagnoses until 2012 obtained from the Danish cancer registry. The associations of HRT with MD and with breast cancer were analyzed separately using Cox's regression. Mediation analyses were used to estimate proportion [with 95% confidence intervals (CI)] of an association between HRT and breast cancer mediated by MD. RESULTS: 2,444 (54.3%) women had mixed/dense breasts, 229 (5.4%) developed breast cancer, and 35.9% were current HRT users at enrollment. Compared to never users, current HRT use was statistically significantly associated with having mixed/dense breasts (relative risk and 95% CI 1.24; 1.14-1.35), and higher risk of breast cancer (hazard ratio 1.87; 1.40-2.48). Association between current HRT use and breast cancer risk was partially mediated by MD (percent mediated = 10%; 95% CI 4-22%). The current HRT use-related breast cancer risk was higher in women with mixed/dense (1.94; 1.37-3.87) than fatty (1.37; 0.80-2.35) breasts (p value for interaction = 0.15). CONCLUSIONS: MD partially mediates some of the association between HRT and breast cancer risk. The association between HRT and breast cancer seems to be stronger in women with dense breasts.

Induction and regulation of murine emphysema by elastin peptides.

Emphysema is the major component of chronic obstructive pulmonary disease (COPD). During emphysema, elastin breakdown in the lung tissue originates from the release of large amounts of elastase by inflammatory cells. Elevated levels of elastin-derived peptides (EP) reflect massive pulmonary elastin breakdown in COPD patients. Only the EP containing the GXXPG conformational motif with a type VIII ß-turn are elastin receptor ligands inducing biological activities. In addition, the COOH-terminal glycine residue of the GXXPG motif seems a prerequisite to the biological activity. In this study, we endotracheally instilled C57BL/6J mice with GXXPG EP and/or COOH-terminal glycine deleted-EP whose sequences were designed by molecular dynamics and docking simulations. We investigated their effect on all criteria associated with the progression of murine emphysema. Bronchoalveolar lavages were recovered to analyze cell profiles by flow cytometry and lungs were prepared to allow morphological and histological analysis by immunostaining and confocal microscopy. We observed that exposure of mice to EP elicited hallmark features of emphysema with inflammatory cell accumulation associated with increased matrix metalloproteinases and desmosine expression and of remodeling of parenchymal tissue. We also identified an inactive COOH-terminal glycine deleted-EP that retains its binding-activity to EBP and that is able to inhibit the in vitro and in vivo activities of emphysema-inducing EP. This study demonstrates that EP are key actors in the development of emphysema and that they represent pharmacological targets for an alternative treatment of emphysema based on the identification of EP analogous antagonists by molecular modeling studies.

Arenophile-Mediated Dearomative Reduction.

A dearomative reduction of simple arenes has been developed which employs a visible-light-mediated cycloaddition of arenes with an N-N-arenophile and in situ diimide reduction. Subsequent cycloreversion or fragmentation of the arenophile moiety affords 1,3-cyclohexadienes or 1,4-diaminocyclohex-2-enes, compounds that are not synthetically accessible using existing dearomatization reactions. Importantly, this strategy also provides numerous opportunities for further derivatization as well as site-selective functionalization of polynuclear arenes.

Long-term exposure to air pollution and mammographic density in the Danish Diet, Cancer and Health cohort.

BACKGROUND: Growing evidence suggests that air pollution may be a risk factor for breast cancer, but the biological mechanism remains unknown. High mammographic density (MD) is one of the strongest predictors and biomarkers of breast cancer risk, but it has yet to be linked to air pollution. We investigated the association between long-term exposure to traffic-related air pollution and MD in a prospective cohort of women 50 years and older. METHODS: For the 4,769 women (3,930 postmenopausal) participants in the Danish Diet, Cancer and Health cohort (1993-1997) who attended mammographic screening in Copenhagen (1993-2001), we used MD assessed at the first screening after cohort entry. MD was defined as mixed/dense or fatty. Traffic-related air pollution at residence was assessed by modeled levels of nitrogen oxides (NOx) and nitrogen dioxide (NO2). The association between mean NOx and NO2 levels since 1971 until cohort baseline (1993-97) and MD was analyzed using logistic regression, adjusting for confounders, and separately by menopause, smoking status, and obesity. RESULTS: We found inverse, statistically borderline significant associations between long-term exposure to air pollution and having mixed/dense MD in our fully adjusted model (OR; 95% CI: 0.96; 0.93-1.01 per 20 µg/m(3) of NOx and 0.89; 0.80- 0.98 per 10 µg/m(3) of NO2). There was no interaction with menopause, smoking, or obesity. CONCLUSION: Traffic-related air pollution exposure does not increase MD, indicating that if air pollution increases breast cancer risk, it is not via MD.

Loss of symmetric cell division of apical neural progenitors drives DENND5A-related developmental and epileptic encephalopathy.

Developmental and epileptic encephalopathies (DEEs) feature altered brain development, developmental delay and seizures, with seizures exacerbating developmental delay. Here we identify a cohort with biallelic variants in DENND5A, encoding a membrane trafficking protein, and develop animal models with phenotypes like the human syndrome. We demonstrate that DENND5A interacts with Pals1/MUPP1, components of the Crumbs apical polarity complex required for symmetrical division of neural progenitor cells. Human induced pluripotent stem cells lacking DENND5A fail to undergo symmetric cell division with an inherent propensity to differentiate into neurons. These phenotypes result from misalignment of the mitotic spindle in apical neural progenitors. Cells lacking DENND5A orient away from the proliferative apical domain surrounding the ventricles, biasing daughter cells towards a more fate-committed state, ultimately shortening the period of neurogenesis. This study provides a mechanism for DENND5A-related DEE that may be generalizable to other developmental conditions and provides variant-specific clinical information for physicians and families.

Loss of symmetric cell division of apical neural progenitors drives DENND5A-related developmental and epileptic encephalopathy.

Developmental and epileptic encephalopathies (DEEs) are a heterogenous group of epilepsies in which altered brain development leads to developmental delay and seizures, with the epileptic activity further negatively impacting neurodevelopment. Identifying the underlying cause of DEEs is essential for progress toward precision therapies. Here we describe a group of individuals with biallelic variants in DENND5A and determine that variant type is correlated with disease severity. We demonstrate that DENND5A interacts with MUPP1 and PALS1, components of the Crumbs apical polarity complex, which is required for both neural progenitor cell identity and the ability of these stem cells to divide symmetrically. Induced pluripotent stem cells lacking DENND5A fail to undergo symmetric cell division during neural induction and have an inherent propensity to differentiate into neurons, and transgenic DENND5A mice, with phenotypes like the human syndrome, have an increased number of neurons in the adult subventricular zone. Disruption of symmetric cell division following loss of DENND5A results from misalignment of the mitotic spindle in apical neural progenitors. A subset of DENND5A is localized to centrosomes, which define the spindle poles during mitosis. Cells lacking DENND5A orient away from the proliferative apical domain surrounding the ventricles, biasing daughter cells towards a more fate-committed state and ultimately shortening the period of neurogenesis. This study provides a mechanism behind DENND5A-related DEE that may be generalizable to other developmental conditions and provides variant-specific clinical information for physicians and families.

Behavioral Changes after Psychiatric Genetic Counseling: An Exploratory Study.

INTRODUCTION: Though it is well established that genetic information does not produce behavior changes, there are limited data regarding whether genetic counseling can facilitate changes in lifestyle and health behaviors that can result in improved health outcomes. METHODS: To explore this issue, we conducted semi-structured interviews with 8 patients who had lived experience of psychiatric illness and who had received psychiatric genetic counseling (PGC). Using interpretive description, we used a constant comparative approach to data analysis. RESULTS: Participants talked about how, prior to PGC, they held misconceptions and/or uncertainties about the causes of and protective behaviors associated with mental illness, which caused feelings of guilt, shame, fear, and hopelessness. Participants reported that PGC reframed things in a way that provided them a sense of agency over illness management, allowed a greater acceptance of illness, and provided release from some of the negative emotions associated with their initial framing of their illness, which seemed to be related to the self-reported increase in engagement in illness management behaviors and consequently improved mental health outcomes. CONCLUSION: This exploratory study provides evidence to support the idea that through addressing emotions associated with perceived cause of illness and facilitating understanding of etiology and risk-reducing strategies, PGC may lead to an increase in behaviors, which protect mental health.

Metastatic Prostate Adenocarcinoma Masquerading as Invasive Fungal Sinusitis.

A 58 year old male with a history of prostate adenocarcinoma presented with diplopia, severe headaches, and eye pain, consistent with sinusitis. Imaging was concerning for invasive fungal sinusitis (IFS) and an urgent ENT consultation was requested. Endoscopic sinus surgery was performed revealing metastatic prostate adenocarcinoma to the sinuses and anterior cranial fossa. The distinctive imaging features in this case are very useful when considering the divergent management options of IFS and metastatic sinus disease. These entities are likely to be encountered more frequently as immunomodulating therapies expand and prostate cancer continues to be a leading cause of death in males.

Health Inequalities in the Use of Telehealth in the United States in the Lens of COVID-19.

The use of remote health care services, or telehealth, is a promising solution for providing health care to those unable to access care in person easily and thus helping to reduce health inequalities. The COVID-19 pandemic and resulting stay-at-home orders in the United States have created an optimal situation for the use of telehealth services for non-life-threatening health care use. A retrospective cohort study was performed using Kantar's Claritis™ database, which links insurance claims encounters (Komodo Health) with patient-reported data (Kantar Health, National Health & Wellness Survey). Logistic regression models (odds ratios [OR], 95% confidence intervals [CI]) examined predictors of telehealth versus in-person encounters. Adults ages ≥18 years eligible for payer-complete health care encounters in both March 2019 and March 2020 were identified (n = 35,376). Telehealth use increased from 0.2% in 2019 to 1.9% in 2020. In adjusted models of respondents with ≥1 health care encounter (n = 11,614), age, marital status, geographic residence (region; urban/rural), and presence of anxiety or depression were significant predictors of telehealth compared with in-person use in March 2020. For example, adults 45-46 years versus 18-44 years were less likely to use telehealth (OR 0.684, 95% CI: 0.561-0.834), and respondents living in urban versus rural areas were more likely to use telehealth (OR 1.543, 95% CI: 1.153-2.067). Substantial increases in telehealth use were observed during the onset of the COVID-19 pandemic in the United States; however, disparities existed. These inequalities represent the baseline landscape that population health management must monitor and address during this pandemic.

Sociodemographic disparities in the management of type 2 diabetes in the United States.

Objective: To examine the potential sociodemographic disparities in type 2 diabetes (T2D) management and care among US adult individuals, after controlling for clinical and behavioral factors.Methods: This was a retrospective cohort study of individuals with T2D (N = 4552) from a linked database of the National Health and Wellness Survey and a large US ambulatory electronic health record (EHR) database. This study period was between 1 January 2015 and 31 December 2018 and individuals were followed up for at least 6 months through EHR after the completion of the survey. The sociodemographic characteristics included gender, race, ethnicity, marital status, education, employment status, household income, insurance status, and geographic region. The independent variables included testing and control of HbA1c, blood pressure (BP), and low-density lipoprotein-cholesterol (LDL-C); hypoglycemia, emergency room (ER) visits, and all-cause hospitalization. Multivariable analyses were conducted using generalized linear models.Results: The percentage of uncontrolled HbA1c was 38.6%. With clinical and behavioral characteristics adjusted, individuals living in the Northeast region had 30% higher odds of having HbA1c testing than those who lived in the South. Blacks and Asians were less likely to have HbA1c control than Whites. Uninsured individuals had a lower likelihood of receiving HbA1c, BP, or LDL-C testing compared with commercial insurers. Individuals with low income were more likely to have higher ER visits and hospitalizations.Conclusion: Potential sociodemographic disparities exist in T2D management and care in the US, indicating the needs for improvement in healthcare access, educational and behavioral programs, as well as disease and treatment management in these subgroups.

Role of the amino acid sequence in domain swapping of the B1 domain of protein G.

Substitution of four amino acid residues (L5V,F30V, Y33F,A34F) in the B1 domain of the immunoglobulin G binding protein (GB1) leads to the formation of a swapped dimer, shown to be in equilibrium with a native-like monomeric state of the protein (Byeon et al., J Mol Biol 2003;333:141-152). In this study, we employ protein design calculations and molecular dynamics simulations to investigate the role of these substitutions in fostering the swapping reaction. DESIGNER, a fully automatic procedure for computing the amino acid sequences likely to stabilize a given backbone structure is used to investigate the effect of the four substitutions on the stability of the wild type native monomeric conformation. Results indicate that at least three of these substitutions (L5V,F30V, A34F) have a destabilizing effect. The L5V forms destabilizing interactions with surrounding residues, whereas F30V causes local strain due to unfavorable interactions with its own backbone. A dual role in the swapping reaction is played by A34F. It destabilizes the monomer conformation while stabilizing the swapped dimer. Our calculations find an energetically favorable conformation for the 34F side chain in the core of the monomer, but only at the expense of forcing the wild type W43 side chain into a highly strained rotamer, and forming unfavorable interactions with both W43 and V54. Although detailed calculations could not be performed on the swapped dimer, due to the lower accuracy of the model, analysis revealed that the 34F side chain from both subunits are tightly packed against each other in the dimer core, suggesting that their replacement by the smaller Ala, as in the wild type, would be highly destabilizing through the creation of a large internal cavity possibly accompanied by a substantial conformational change. Analysis of room temperature molecular dynamics (MD) simulations of the wild type and the modeled quadruple mutant structures reveals that the latter structure fluctuates more than its wild type counterpart. In addition, its C-terminal beta-hairpin, which is exchanged in the swapping reaction, undergoes a conformation change, which pushes it further away from the remainder of the protein. Simulations at higher temperature (450 K) show that the quadruple mutant unfolds earlier and more completely than the wild type following a sequence of events that is compatible with the description of the highly fluctuating monomeric state of this mutant observed by NMR. Our findings thus support the notion that domain swapping in GB1 is fostered by three main factors: a decrease in stability and increased flexibility of the monomer conformation, concomitant with stabilization of the swapped dimer conformation through new interactions that have no counterparts in the monomer.