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Core Histones Are Constituents of the Perinuclear Theca of Murid

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Core Histones Are Constituents of the Perinuclear Theca of Murid International Journal of Molecular Sciences Article Core Histones Are Constituents of the Perinuclear Theca of Murid Spermatozoa: An Assessment of Their Synthesis and Assembly during Spermiogenesis and Function after Gametic Fusion Lauren E. Hamilton 1,2, Morgan Lion 1, Luis Aguila 3 , João Suzuki 3, Genevieve Acteau 1, Nicole Protopapas 1, Wei Xu 1, Peter Sutovsky 2,4 , Mark Baker 5 and Richard Oko 1,* 1 Department of Biomedical and Molecular Sciences, Queen’s University, Botterell Hall, 18 Stuart Street, Kingston, ON K7L 3N6, Canada; [email protected] (L.E.H.); [email protected] (M.L.); [email protected] (G.A.); [email protected] (N.P.); [email protected] (W.X.) 2 Division of Animal Science, College of Agriculture, Food and Natural Resources, University of Missouri, Columbia, MO 65211, USA; [email protected] 3 Center for Research in Reproduction and Fertility, Department of Veterinary Sciences, Université de Montreal, St. Hyacinthe, QC J2S 2M2, Canada; [email protected] (L.A.); [email protected] (J.S.) 4 Department of Obstetrics, Gynecology and Women’s Health, School of Medicine, University of Missouri, Columbia, MO 65211, USA Citation: Hamilton, L.E.; Lion, M.; 5 School of Environmental and Life Science, University of Newcastle, Callaghan, NSW 2308, Australia; Aguila, L.; Suzuki, J.; Acteau, G.; [email protected] Protopapas, N.; Xu, W.; Sutovsky, P.; * Correspondence: [email protected] Baker, M.; Oko, R. Core Histones Are Constituents of the Perinuclear Theca Abstract: The perinuclear theca (PT) of the eutherian sperm head is a cytoskeletal-like structure that of Murid Spermatozoa: An houses proteins involved in important cellular processes during spermiogenesis and fertilization. Assessment of Their Synthesis and Building upon our novel discovery of non-nuclear histones in the bovine PT, we sought to investigate Assembly during Spermiogenesis and whether this PT localization was a conserved feature of eutherian sperm. Employing cell fractionation, Function after Gametic Fusion. Int. J. immunodetection, mass spectrometry, qPCR, and intracytoplasmic sperm injections (ICSI), we Mol. Sci. 2021, 22, 8119. https:// doi.org/10.3390/ijms22158119 examined the localization, developmental origin, and functional potential of histones from the murid PT. Immunodetection localized histones to the post-acrosomal sheath (PAS) and the perforatorium Academic Editors: (PERF) of the PT but showed an absence in the sperm nucleus. MS/MS analysis of selectively Katerina Komrskova, extracted PT histones indicated that predominately core histones (i.e., H3, H3.3, H2B, H2A, H2AX, Pavla Postlerova and and H4) populate the murid PT. These core histones appear to be de novo-synthesized in round Michaela Frolikova spermatids and assembled via the manchette during spermatid elongation. Mouse ICSI results suggest that early embryonic development is delayed in the absence of PT-derived core histones. Received: 5 July 2021 Here, we provide evidence that core histones are de novo-synthesized prior to PT assembly and Accepted: 26 July 2021 deposited in PT sub-compartments for subsequent involvement in chromatin remodeling of the male Published: 29 July 2021 pronucleus post-fertilization. Publisher’s Note: MDPI stays neutral Keywords: histones; gamete biology; spermiogenesis; microtubular manchette; spermatozoa; perinu- with regard to jurisdictional claims in clear theca; postacrosomal sheath; perforatorium; fertilization; ICSI; mass spectrometry published maps and institutional affil- iations. 1. Introduction There are three main compartments within the sperm head: the nucleus, the acrosome, Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. and the perinuclear theca (PT) (Figure1). The acrosome is a secretory vesicle that encapsu- This article is an open access article lates the anterior region of the head and contains proteins required for oocyte penetration. distributed under the terms and Underneath the acrosome and surrounding the nucleus is the PT, a condensed cytosolic- conditions of the Creative Commons protein-rich layer [1]. Based on compositional and functional considerations, the PT can Attribution (CC BY) license (https:// be divided into two compartments: the subacrosomal layer (SAL), which houses proteins creativecommons.org/licenses/by/ involved in the assembly and attachment of the acrosome to the spermatid nucleus during 4.0/). Int. J. Mol. Sci. 2021, 22, 8119. https://doi.org/10.3390/ijms22158119 https://www.mdpi.com/journal/ijms Int. J. Mol. Sci. 2021, 22, x FOR PEER REVIEW 2 of 25 Int. J. Mol. Sci. 2021, 22, 8119 2 of 26 considerations, the PT can be divided into two compartments: the subacrosomal layer (SAL), which houses proteins involved in the assembly and attachment of the acrosome to the spermatid nucleus during spermiogenesis [1–6], and the post-acrosomal sheath (PAS),spermiogenesis which houses [1–6], proteins and the post-acrosomalimplicated in oocyte sheath activation (PAS), which and houses zygote proteins development impli- duringcated in and oocyte after activation fertilization and zygote [1,7–14]. development In murids, during the andPT has after an fertilization additional[1 ,7third–14]. compartmentIn murids, the termed PT has the an additionalperforatorium third [15] compartment (Figure 1). termedThe perforatorium the perforatorium is a hook- [15] shaped(Figure 1extension). The perforatorium of the PT unique is a hook-shaped to falciform extensionspermatozoa of the that PT encircles unique toand falciform extends spermatozoa that encircles and extends beyond the apical nucleus. It is compositionally beyond the apical nucleus. It is compositionally distinct from the SAL but has distinct from the SAL but has developmental, compositional, and functional similarities developmental, compositional, and functional similarities with the PAS [15]. with the PAS [15]. Figure 1. Diagrammatic representations of mid-sagittal sections through falciform (A) rat and (B) Figuremouse sperm1. Diagrammatic heads and spatulaterepresentations (C) bull of sperm mid-sa heads.gittal Regardlesssections through of morphological falciform (A variations,) rat and the(B) mouse sperm heads and spatulate (C) bull sperm heads. Regardless of morphological variations, eutherian sperm head is comprised of three major cellular compartments: the nucleus (N; black), the eutherian sperm head is comprised of three major cellular compartments: the nucleus (N; black), the acrosome (A; crosshatched pattern), and the perinuclear theca (PT; red/yellow/orange). The the acrosome (A; crosshatched pattern), and the perinuclear theca (PT; red/yellow/orange). The nucleusnucleus occupies occupies the the majority of the specialized sperm head volume. The acrosome overlays the apicalapical half half of of the nucleus and is bound by thethe inner and outer acrosomal membranes (IAM and OAM,OAM, respectively).respectively). InIn the the mid-sagittal mid-sagittal sections sections of the of muridthe murid sperm sperm (A,B), the(A, areaB), the that area the acrosomethat the acrosomecovers is superimposedcovers is superimposed over each sectionover each for sect reference.ion for Thereference. PT is formedThe PT byis severalformed continuousby several continuouscytosolic compartments, cytosolic compartments, which together which encapsulate together the encapsulate entire condensed the enti nucleus,re condensed except where nucleus, the excepttail implants. where the The tail PT implants. of spatulate The sperm PT of (spatulateC) is subdivided sperm (C into) is thesubdivided sub-acrosomal into the layer sub-acrosomal (SAL; red) layerand the (SAL; post-acrosomal red) and the sheath post-acrosomal (PAS; yellow). sheath The (P equatorialAS; yellow). segment The equatorial (ES) is the segment thin caudal (ES) portion is the thin caudal portion of the acrosome, covered on its outer surface by an extension of the SAL, the of the acrosome, covered on its outer surface by an extension of the SAL, the outer periacrosomal outer periacrosomal layer (OPL). In addition to SAL, OPL, and PAS regions, the PT of falciform layer (OPL). In addition to SAL, OPL, and PAS regions, the PT of falciform sperm contains the sperm contains the perforatorium (orange), which extends apically from the nucleus to form the characteristicperforatorium hook (orange), shape. which The sperm extends tail apicallyis partially from shown the nucleusextending to from form the the caudal characteristic nucleus. hook The coreshape. structure, The sperm the axoneme tail is partially (AX), is shownsurrounded extending by outer from dense the fibe caudalrs, which nucleus. in turn The are core surrounded structure, bythe helically axoneme arranged (AX), is mitochondria surrounded by (M) outer in the dense midpiece fibers, of which the tail. in turnThe plasmalemma are surrounded (PM) by helically encases thearranged sperm mitochondriahead and tail. (M)Labels: in the HCS, midpiece displaced ofthe head tail. cap The segment: plasmalemma VS, ventral (PM) spur. encases Adapted the sperm from Protopapashead and tail. et Labels:al. [15], HCS,Oko and displaced Marave headi [16], cap and segment: Lalli and VS, Clermont ventral spur. [17].Adapted from Protopapas et al. [15], Oko and Maravei [16], and Lalli and Clermont [17]. The fraction of histones retained in the mature eutherian spermatozoon nucleus is species-dependent.The fraction of The histones amount retained of histones in the foun matured in the eutherian spermatid spermatozoon nucleus to date nucleus is also is highlyspecies-dependent.
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