Biology subjects

Palm, G.

Publications and source records attributed to Palm, G..

5 recordsLinked to original sources

bioRxiv2026

Structure of the Pre-Initiation Complex Explains CMGE Biogenesis

Summary paragraphWhen cells enter S phase, bidirectional DNA replication is initiated through the kinase-regulated recruitment of three activators (Cdc45, GINS and Pol epsilon) to a duplex DNA-loaded double hexamer of MCM ATPases. Together these proteins form two CMGE helicases that establish divergent replication forks as they become separated1. To understand CMGE biogenesis, we reconstituted the pre-Initiation Complex with purified yeast proteins. The cryo-EM structure shows a set of firing factors caught in the act of assembling two symmetric CMGEs. We show how stepwise complex formation reshapes MCM in preparation for DNA opening and we explain how ATP promotes firing-factor ejection and CMGE maturation. While we find that Sld2 promotes GINS recruitment to MCM as expected, it also aids efficient separation of the CMGE dimer, and it is essential for lagging strand ejection from MCM. These findings have direct implications for our understanding of the metazoan Sld2 ortholog, RECQL4, pointing to a replication-fork establishment mechanism conserved across eukaryotes.

biochemistry↗

bioRxiv2026

Mcm10 braces converging replicative helicases to pull apart origin DNA

Eukaryotic DNA replication initiation requires controlled, temporally separated steps to preserve genome stability. The MCM replicative helicase is loaded on duplex DNA as an inactive double hexamer, which nucleates the assembly of dimeric Cdc45-MCM-GINS-Pol epsilon (dCMGE) replisomes. Mcm10 splits dCMGE into two, generating divergent replication forks, but the mechanism is unknown. Using ATPase-defective yeast MCM variants that slow origin melting, we captured five reaction intermediates that explain the structural mechanism. Two Mcm10 molecules bridge across the CMGE dimer, bracing two converging helicases. The restrained MCM motors pull apart the two DNA filaments, such that each lagging strand becomes ejected through the Mcm2-5 gate. Our reconstituted structures resemble the double CMG stabilized by metazoan DONSON, pointing to an origin DNA melting mechanism conserved across evolution.

biochemistry↗

bioRxiv2026

A Quality Measure for Repeating Multiple-Unit Spike Patterns

We propose a quality measure for spatio-temporal spike patterns (STPs) in multiple-neuron recordings. In such recordings, repeating STPs or pattern repetitions (PRs) are often found, with many of these generated by chance. To rule those out, statistical tests have been developed to discriminate the unlikely from the more likely PRs. This statistical problem is complicated by the fact that there are several obvious quality criteria for a PR, such as the size (the number of spikes) of the pattern and the number of its occurrences. Here, we propose a canonical way of combining several criteria (which we collect in the so-called signature of the pattern) into a single quality measure, based on the unlikeliness of the pattern. This measure is defined mathematically, and a formula for its computation is derived for stationary spike trains. It can be used to compare PRs. Since spike trains are not stationary in practice, we discuss, for two experimental data sets, how well the stationary formula correlates with the defined quality measure as determined from simulations. The results encourage the use of the stationary formula or also some simpler, related formulas as proxies for the quality, for the comparison of PRs and also for statistical tests that avoid the multiple testing problem incurred by using several quality criteria. Based on our results, we propose a few test statistics, i.e., random variables on the space of multi-unit spike trains with an appropriate null-hypothesis distribution, to evaluate STPs with less computational and sampling efforts.

neuroscience↗

bioRxiv2022

Molecular genetics of GLUT1DS Italian pediatric cohort: 10 novel related-disease variants and structural analysis

GLUT1 deficiency syndrome (GLUT1DS1; OMIM #606777) is a rare genetic metabolic disease, characterized by infantile-onset epileptic encephalopathy, global developmental delay, progressive microcephaly and movement disorders (e.g. spasticity and dystonia). It is caused by heterozygous mutations in the SLC2A1 gene, which encodes the GLUT1 protein, a glucose transporter across the blood-brain barrier (BBB). Most commonly these variants arise de novo resulting in sporadic cases, although several familial cases with AD inheritance pattern have been described. Twenty-seven Italian pediatric patients clinically suspect of GLUT1DS from both sporadic and familial cases have been enrolled. We detected by trios sequencing analysis 25 different variants causing GLUT1DS. Of these, 40% of the identified variants (10 out of 25) had never been reported before, including missense, frameshift and splice site variants. Their X-ray structure analyses strongly suggested the potential pathogenic effects of these novel disease-related mutations, broadening the genotypic spectrum heterogeneity found in the SLC2A1 gene. Moreover, 24% is located in a vulnerable region of the GLUT1 protein that involves transmembrane 4 and 5 helices encoded by exon 4, confirming a mutational hotspot in the SLC2A1 gene. Lastly, we investigated possible correlations between mutation type and clinical and biochemical data observed in our GLUT1DS cohort, revealing that splice site and frameshift variants are related to a more severe phenotype and low CSF parameters. Author summaryWe investigated the molecular data of 27 pediatric patients clinically suspect of GLUT1 deficiency syndrome. By performing trios sequencing analysis, we highlighted ten novel disease-related variants, and their X-ray structure analyses, suggesting the pathogenic effects of these identified mutations. Moreover, the wide clinical and genetic heterogeneity observed in our cohort allowed possible correlations between mutation type and clinical and biochemical data. This analysis enabled to delineate that splice site and frameshift variants are related to a more severe phenotype and low CSF/glucose values. Further clinical and genetic/epigenetics studies could clearly the high phenotypic variability observed in these patients.

genetics↗

bioRxiv2021

Generating surrogates for significance estimation ofspatio-temporal spike patterns

The generation of surrogate data, i.e., the modification of original data to destroy a certain feature, is used for the implementation of a null-hypothesis whenever an analytical approach is not feasible. Thus, surrogate data generation has been extensively used to assess the significance of spike correlations in parallel spike trains. In this context, one of the main challenges is to properly construct the desired null-hypothesis distribution and to avoid a bias in the null-hypothesis by altering the spike train statistics. A classical surrogate technique is uniform dithering (UD), which displaces spikes locally and uniformly. In this study, we compare UD against five surrogate techniques (two newly introduced) in the context of the detection of significant spatio-temporal spike patterns. We evaluate the surrogates for their performance, first on spike trains based on point process models with constant firing rate, and second on modeled non-stationary artificial data serving as ground truth to assess the pattern detection in a more complex and realistic setting. We determine which statistical features of the original spike trains are modified and to which extent. Moreover, we find that UD fails as an appropriate surrogate because it leads to a loss of spikes in the context of binning and clipping, and thus to a large number of false-positive patterns. The other surrogates achieve a better performance in detecting precisely timed higher-order correlations. Based on these insights, we analyze experimental data from pre-/motor cortex of macaque monkeys during a reaching-and-grasping task for spatio-temporal spike patterns. Significance statementTemporal jittering or dithering of single spikes or subsections of spike trains is a common method of generating surrogate data for the statistical analysis of temporal spike correlations. We discovered a serious problem with the classical and widely used method of uniform dithering that can lead to an overestimation of significance, i.e., to false positives in the statistical evaluation of spatio-temporal spike patterns. Therefore we consider 5 other dithering methods, compare and evaluate their statistical properties. Finally, we apply a much better method (trial shifting) to the analysis of experimental multiple-unit recordings and find several highly significant patterns that also reflect different experimental situations.

neuroscience↗