Biology subjects

Pazos, M.

Publications and source records attributed to Pazos, M..

6 recordsLinked to original sources

bioRxiv2025

Targeting Monoallelic CREBBP/EP300 Mutations in Germinal Center-Derived B-Cell Lymphoma with a First-in-Class Histone Acetyltransferase Activator

Germinal center B-cell lymphomas frequently exhibit monoallelic loss-of-function mutations in the histone acetyltransferases CREBBP and p300, which contribute to lymphoma development, disrupt normal germinal center biology, and promote immune evasion. This study evaluates YF2, a histone acetyltransferase activator that increases CREBBP/p300 activity, as a potential therapeutic approach for B-cell lymphoma. YF2 binds the bromodomain of CREBBP/p300, increasing their auto-acetylation and enzymatic activity. It also induces cytotoxicity in B-cell lymphoma cell lines, with a stronger response in those carrying CREBBP/EP300 mutations. Additionally, YF2 increases the acetylation of key CREBBP/p300 substrates, including H3K27, p53, and BCL6, leading to enhanced apoptosis and altered B-cell diTerentiation. YF2 is well tolerated in vivo and extends survival in both cell line- and patient-derived xenograft mouse models. Moreover, YF2 exposure upregulates antigen-presentation markers and reshapes the immune microenvironment, strengthening responses to immune checkpoint blockade. Taken together, these findings support pharmacologic activation of CREBBP/p300 with YF2 as a compelling therapeutic strategy for B-cell lymphoma. SignificanceYF2 stimulates CREBBP/p300 activity to normalize B-cell diTerentiation and enhance immune recognition, oTering a potential approach to limit lymphoma progression and strengthen immunotherapy responses.

cancer biology↗

bioRxiv2025

A novel peptidoglycan deacetylase modulates daughter cell separation in E. coli

Peptidoglycan hydrolases facilitate bacterial cell wall growth by creating space for insertion of new material and allowing physical separation of daughter cells. In Escherichia coli, three peptidoglycan amidases, AmiA, AmiB and AmiC, cleave septal peptidoglycan during cell division. The LytM-domain proteins EnvC and NlpD activate these amidases either from inside the cell or the outer membrane: EnvC binds to the cytoplasmic membrane-anchored divisome components FtsEX, and NlpD and ActS are outer membrane lipoproteins. Here we report the identification of a novel periplasmic deacetylase called SddA that removes acetyl groups from denuded peptidoglycan glycan strands, the products of amidases. SddA is a substrate for the periplasmic protease Prc, suggesting regulation via protein degradation. The sddA gene is co-expressed with the gene encoding EnvC, linking SddA function to amidase activation. Consistent with this link, the deletion of sddA alleviates phenotypes associated with lack of amidase activation, while overexpression of sddA alleviates phenotypes related to a defective Tol-Pal system and causes cell chaining due to reduced septum peptidoglycan cleavage unless envC is co-expressed. We present a model according to which SddA modulates the activation of the septum-splitting amidases during cell division. AUTHOR SUMMARYBacteria surround their cell membrane by the essential peptidoglycan (cell wall) layer to prevent bursting open due to their turgor. During cell division, bacteria produce a septum at midcell, which must be cleaved for daughter cells to separate. Here, we report the identification of a new enzyme, SddA, that modifies a particular type of peptidoglycan material, denuded glycan chains released during the splitting of septal peptidoglycan for daughter cell separation, in the Gram-negative Escherichia coli. We propose a model in which SddA modulates a switch in the septal peptidoglycan splitting, ensuring splitting is activated from the cell membrane in the early stages of cell division and from the outer membrane in the late stages.

microbiology↗

bioRxiv2022

Peptidoglycan maturation controls spatiotemporal organisation of outer membrane proteins in Escherichia coli

Linkages between the outer membrane of Gram-negative bacteria and the peptidoglycan layer are crucial to the maintenance of cellular integrity and enable survival in challenging environments1-5. The functionality of the outer membrane relies on outer membrane proteins (OMPs), which are inserted by the {beta}-barrel assembly machine, BAM6, 7. Previous work has shown that growing Escherichia coli cells segregate old OMPs towards the poles by an unknown mechanism8. Here, we demonstrate that peptidoglycan underpins the spatiotemporal organisation of OMPs. Mature, tetrapeptide-rich peptidoglycan binds to BAM components and suppresses OMP foldase activity. Nascent peptidoglycan, which is enriched in pentapeptides and concentrated at septa9, associates with BAM poorly and has little impact on its activity, leading to preferential insertion of OMPs at division sites. Synchronising OMP biogenesis to cell wall growth enables bacteria to replenish their OMPs by binary partitioning. Our study reveals that Gram-negative bacteria coordinate the assembly of two major cell envelope layers by rendering OMP biogenesis responsive to peptidoglycan maturation. This coordination offers new possibilities for the design of antibiotics that disrupt cell envelope integrity.

microbiology↗

bioRxiv2021

PBP1 of Staphylococcus aureus has multiple essential functions in cell division

Bacterial cell division is a complex process requiring the coordination of multiple components, to allow the appropriate spatial and temporal control of septum formation and cell scission. Peptidoglycan (PG) is the major structural component of the septum, and our recent studies in the human pathogen Staphylococcus aureus have revealed a complex, multi- stage PG architecture that develops during septation. Penicillin binding proteins (PBPs) are essential for the final steps of PG biosynthesis - their transpeptidase activity links together the peptide sidechain of nascent glycan strands together. PBP1 is required for cell division in S. aureus and here we demonstrate that it has multiple essential functions associated with its enzymatic activity and as a regulator of division. Loss of PBP1, or just its C-terminal PASTA domains, results in cessation of division at the point of septal plate formation. The PASTA domains can bind PG and thus coordinate the cell division process. The transpeptidase activity of PBP1 is also essential but its loss leads to a strikingly different phenotype of thickened and aberrant septa, which is phenocopied by the morphological effects of adding the PBP1-specific {beta}-lactam, meropenem. Together these results lead to a model for septal PG synthesis where PBP1 enzyme activity is responsible for the characteristic architecture of the septum and PBP1 protein molecules coordinate cell division allowing septal plate formation.

microbiology↗

bioRxiv2019

The outer membrane lipoprotein NlpI nucleates hydrolases within peptidoglycan multi-enzyme complexes in Escherichia coli

The peptidoglycan (PG) sacculus provides bacteria with the mechanical strength to maintain cell shape and resist osmotic stress. Enlargement of the mesh-like sacculus requires the combined activity of PG synthases and hydrolases. In Escherichia coli, the activity of the two bifunctional PG synthases is driven by lipoproteins anchored in the outer membrane. However, the regulation of PG hydrolases is less well understood, with only regulators for PG amidases having been described. Here, we identify the lipoprotein NlpI as a general adaptor protein for PG hydrolases. NlpI binds to different classes of hydrolases and can specifically form multimeric complexes with various PG endopeptidases. In addition, NlpI seems to contribute both to PG elongation and cell division biosynthetic complexes based on its localization and genetic interactions. In line with such a role, we reconstitute PG multi-enzyme complexes containing NlpI, the PG synthesis regulator LpoA, its cognate bifunctional synthase, PBP1A, and different endopeptidases. Our results indicate that PG regulators and adaptors are part of PG biosynthetic multi-enzyme complexes, regulating and potentially coordinating the spatiotemporal action of PG synthases and hydrolases.\n\nSignificanceThe activity of PG hydrolases may cause lysis of the bacterial cell if left unregulated. Hence, the cell must have ways of regulating and coordinating their activities. Our current understanding of how this occurs is incomplete. In this work, we present the outer membrane (OM) anchored lipoprotein, NlpI, as a scaffold of peptidoglycan hydrolases. We propose that NlpI facilitates the formation of multi-enzyme complexes and that, along with other regulators, it coordinates a safe enlargement and separation of the PG layer in E. coli.

microbiology↗

bioRxiv2018

Inactivation of the monofunctional peptidoglycan glycosyltransferase SgtB allows Staphylococcus aureus to survive in the absence of lipoteichoic acid

The cell wall of Staphylococcus aureus is composed of peptidoglycan and the anionic polymers lipoteichoic acid (LTA) and wall teichoic acid. LTA is required for growth and normal cell morphology in S. aureus. Strains lacking LTA are usually only viable when grown under osmotically stabilizing conditions or after the acquisition of compensatory mutations. LTA negative suppressor strains with inactivating mutations in gdpP, resulting in an increase in intracellular c-di-AMP levels, have been described previously. Here, we sought to identify factors other than c-di-AMP that allow S. aureus to survive without LTA. LTA-negative strains able to grow in un-supplemented medium were obtained and found to contain mutations in sgtB, mazE, clpX or vraT. The growth improvement through mutations in mazE and sgtB was confirmed by complementation analysis. We also show that an S. aureus sgtB transposon mutant, inactivated for the monofunctional peptidoglycan glycosyltransferase SgtB, displays a 4-fold increase in the MIC towards a number of cell wall-targeting antibiotics, suggesting that alteration in the peptidoglycan structure could help bacteria compensate for the lack of LTA. Muropeptide analysis of peptidoglycan isolated from a WT and sgtB mutant strains did not reveal any sizable alternations in the peptidoglycan structure. In contrast, the peptidoglycan isolated from an LTA-negative ltaS mutant strain showed a significant reduction in the fraction of highly crosslinked peptidoglycan, which was partially rescued in the sgtB/ltaS double mutant suppressor strain. Taken together, these data point towards an important function of LTA in cell wall integrity through its requirement for proper peptidoglycan assembly.\n\nImportanceThe bacterial cell wall acts as primary defence against environmental insults such as changes in osmolarity. It is also a vulnerable structure as defects in its synthesis can lead to growth arrest or cell death. The important human pathogen Staphylococcus aureus has a typical Gram-positive cell wall, which consists of peptidoglycan and the anionic polymers lipoteichoic acid (LTA) and wall teichoic acid. Several clinically relevant antibiotics inhibit the synthesis of peptidoglycan; hence it and teichoic acids are considered attractive targets for the development of new antimicrobials. We show that LTA is required for efficient peptidoglycan crosslinking in S. aureus and inactivation of a peptidoglycan glycosyltransferase can partially rescue this defect, altogether revealing an intimate link between peptidoglycan and LTA synthesis.

microbiology↗