cells Review Multifunctionality of the Telomere-Capping Shelterin Complex Explained by Variations in Its Protein Composition Claire Ghilain 1, Eric Gilson 1,2,3,* and Marie-Josèphe Giraud-Panis 1,* 1 Université Côte d’Azur, CNRS, INSERM, IRCAN, 06000 Nice, France; [email protected] 2 International Research Laboratory for Cancer, Aging and Hematology, Shanghai Ruijin Hospital, Shanghai Jiaotong University and Côte-d’Azur University, Shanghai 200025, China 3 Department of Genetics, CHU Nice, 06000 Nice, France * Correspondence: [email protected] (E.G.); [email protected] (M.-J.G.-P.) Abstract: Protecting telomere from the DNA damage response is essential to avoid the entry into cellular senescence and organismal aging. The progressive telomere DNA shortening in dividing somatic cells, programmed during development, leads to critically short telomeres that trigger replicative senescence and thereby contribute to aging. In several organisms, including mammals, telomeres are protected by a protein complex named Shelterin that counteract at various levels the DNA damage response at chromosome ends through the specific function of each of its subunits. The changes in Shelterin structure and function during development and aging is thus an intense area of research. Here, we review our knowledge on the existence of several Shelterin subcomplexes and the functional independence between them. This leads us to discuss the possibility that the multifunctionality of the Shelterin complex is determined by the formation of different subcomplexes Citation: Ghilain, C.; Gilson, E.; whose composition may change during aging. Giraud-Panis, M.-J. Multifunctionality of the Keywords: telomere; aging; Shelterin; senescence; DNA damage response Telomere-Capping Shelterin Complex Explained by Variations in Its Protein Composition. Cells 2021, 10, 1753. https://doi.org/10.3390/cells10071753 1. Foreword The linear nature of eukaryotic chromosomes causes two serious threats to genome Academic Editors: Christoph Englert, integrity. The first threat stems from DNA extremities, which can be misidentified as DNA Holger Bierhoff and Alexander damage by the DNA damage response (DDR) machinery, leading to cellular senescence, E. Kalyuzhny apoptosis or double-strand break (DSB) repair [1]. The second threat stems from the inabil- ity of the conventional replication machinery to fully replicate the extremities of parental Received: 28 May 2021 DNA. In somatic cells, this leads to an inexorable erosion of chromosomes ends, which is Accepted: 9 July 2021 Published: 11 July 2021 compensated for by activating pathways that replenish telomeres (Telomerase or Alterna- tive Lengthening of Telomeres (ALT), based on recombination) in germ, stem and cancer Publisher’s Note: MDPI stays neutral cells [2]. In addition to this replicative attrition, telomeres are sensitive to a wide range of with regard to jurisdictional claims in endogenous and environmental factors, such as improper cell cycle progression through published maps and institutional affil- mitosis, oxidative or genotoxic stress, alcohol, caffeine, heat shock, stress hormones and iations. psychological stress [3]; therefore, telomeres have emerged as important cell-cycle and senescence regulators, stress sensors, and lifespan predictors. Indeed, experimental and pathological evidence of genome-wide deleterious consequences of telomeres dysfunction are numerous. For example, several premature aging syndromes generically called telom- eropathies originate from or are associated with mutations in telomere-associated proteins Copyright: © 2021 by the authors. Dyskeratosis congenita Licensee MDPI, Basel, Switzerland. ( , Hoyeraal–Hreidarsson, Revesz or Coats plus syndromes, amongst This article is an open access article others) [4]. Furthermore, replenishing telomeres is an obligatory step for oncogenesis. distributed under the terms and Failure to do so causes rampant genome instability (multiple genome rearrangements conditions of the Creative Commons through cycles of break–fusion–breaks, kataegis or chromothripsis) and cell death [5]; thus, Attribution (CC BY) license (https:// the obvious importance of telomeric actors has elicited various therapeutic trials. Although creativecommons.org/licenses/by/ Telomerase could be perceived as a prime target, limitations in Telomerase-based strategies, 4.0/). such as the consequential activation of ALT-driven telomere elongation or the possible Cells 2021, 10, 1753. https://doi.org/10.3390/cells10071753 https://www.mdpi.com/journal/cells Cells 2021, 10, x 2 of 18 Cells 2021, 10, 1753 2 of 18 a prime target, limitations in Telomerase-based strategies, such as the consequential activation of ALT-driven telomere elongation or the possible pro-aging side effects, have pro-agingstimulated side the effects,development have stimulatedof alternative the a developmentpproaches [6]. of Targeting alternative the approachescomplexes that [6]. Targetingform telomeres the complexes themselves that formcould, telomeres thus, be themselves a complementary could, thus, or alternative be a complementary route for ortelomere-based alternative route therapies. for telomere-based therapies. WeWe areare soon soon reaching reaching the the 30 years30 years benchmark benchmark since since the discoverythe discovery of the of first the mam- first malianmammalian telomeric telomeric protein, protein, TRF1 [TRF17–9]. Through[7–9]. Through the years, the 5years, other 5 proteins other proteins (TRF2, RAP1,(TRF2, TIN2,RAP1, TPP1 TIN2, and TPP1 POT1) and have POT1) also beenhave identifiedalso been asidentified being essential as being for theessential protection for the of theseprotection natural of DNA these ends natural and DNA have beenends proposedand have tobeen form proposed a whole complexto form a named whole Shelterin complex bynamed de Lange Shelterin in 2005 by [de10 ]Lange (Figure in1 2005). [10] (Figure 1). Figure 1. The Shelterin complex protects telomere from illegitimate activation of DNA damage Figure 1. The Shelterin complex protects telomere from illegitimate activation of DNA damage responseresponse (ATM/CHK2, (ATM/CHK2, ATR/CHK1) ATR/CHK1) andand repair repair (homology-directed (homology-directed DNA DNA repair, repair, HdDR, HdDR, NHEJ, NHEJ, HR). 2.HR). The Shelterin Complex: Composition and Biological Role 2. TheThe Shelterin Shelterin Complex: ultimate roleComposition is to maintain and Biological telomeres homeostasis,Role which as we have seen, is critical for genome stability and cell fate. Moreover, telomeric proteins participate The Shelterin ultimate role is to maintain telomeres homeostasis, which as we have in several cellular processes not directly related to telomere homeostasis, such as replica- seen, is critical for genome stability and cell fate. Moreover, telomeric proteins participate tion, mitosis, meiosis, heterochromatin stability, immunity or neuronal development [11]; hence,in several the Shelterin cellular complexprocesses is highlynot directly multifunctional related to and telomere plays pivotal homeostasis, roles in telomeresuch as protection,replication, genome mitosis, stability meiosis, and cellheterochromatin fate. Remarkably, stability, each Shelterin immunity subunit or has neuronal specific functionsdevelopment in these [11]; processes, hence, the leading Shelterin us tocomp questionlex is thehighly respective multifunctional contributions and of plays the wholepivotal complex roles in versus telomere subcomplexes protection, and genome whether stability the protein and compositioncell fate. Remarkably, of the Shelterin each complexShelterin changes subunit during has specific aging. functions in these processes, leading us to question the respectiveOn a molecular contributions level, of this the complex whole comp is vitallex for versus efficient subcomplexes replication ofand the whether G-rich and the repetitiveprotein composition telomeric DNA of the [ 12Shelterin–14] and complex to regulate changes Telomerase- during aging. or ALT-driven elongation of telomeres.On a molecular Importantly, level, thethis six complex telomeric is vital proteins for efficient protect replication chromosome of endsthe G-rich from theand DNArepetitive damage telomeric response DNA (through [12–14] inhibitionand to regu oflate the Telomerase- ataxia–telangiectasia or ALT-driven mutated elongation kinase (ATM)of telomeres. and the Importantly, ataxia–telangiectasia the six telomeric and Rad3-related proteins protect kinase (ATR)chromosome pathways) ends and from from the repairDNA machineriesdamage response (non-homologous (through inhibition end joining of the (NHEJ) ataxia–telangiectasia and homologous mutated recombination kinase (HR)) [15]. Cells 2021, 10, 1753 3 of 18 Biochemical and structural studies have revealed interactions between sets of these proteins that have led to the definition of the complex containing the 6 proteins. The concept of Shelterin was not built in one day. It took more than 10 years from the identification of TRF1 by de Lange’s laboratory in the early 1990s [8] until the publication of TPP1 in 2004 simultaneously by three different groups [16]; however, deciphering the role of each of the members took decades, and even to this day new connections and functions of these proteins are being discovered. One crucial notion concerning the Shelterin proteins is their roles as hubs: recruiting proteins, regulating activities and controlling conformation of many different molecules, the first one being telomeric DNA. Binding telomeric