Importance and Meaning of Phenoptosis Giacinto Libertini M.D., Independent Researcher,

1. Importance and Meaning of Phenoptosis Giacinto Libertini M.D., Independent Researcher, External collaborator of Dept. of Translational Medical Sciences, Federico II Univ., Naples, Italy www.r-site.org/ageing e-mail: giacinto.libertini@tin.it

2. The neologism “phenoptosis”, coined in analogy with “apoptosis”, was proposed in 1997 by Skulachev, an authoritative biochemist, to indicate the “programmed death of an organism”[1]. In 2010, in a workshop, Skulachev proposed a yearly special issue of the journal directed by him, Biochemistry (Moscow), dedicated to Phenoptosis [2]. In the first number of these issues, the definition was extended: “Phenoptosis is the death of an individual, caused by its own actions or by actions of close relatives*, and not caused primarily by accidents or diseases or external factors, which is determined, regulated or influenced by genes favoured by natural selection.” [3] * siblicide; in particular, the parent-caused death of an offspring or filial infanticide. [1] Skulachev VP (1997) Aging is a specific biological function rather than the result of a disorder in complex living systems: biochemical evidence in support of Weismann’s hypothesis.Biochem (Mosc) 62:1191-5 [2] Skulachev VP (2010, May 26) The talk at the "From Homo sapiens to Homo sapiens liberatus" workshop, Moscow (Russia) [3] Libertini G (2012) Classification of Phenoptotic Phenomena. Biochem (Mosc) 77:707-15 2/20

3. Phenoptosis, alias phenoptotic phenomena, may be classified as [1]: direct (the organism kills itself) P h e no p t o sis indirect (the organism kills a close relative) optional (conditional) rapid (e.g., semelparity) obligatory slow (aging) determined by biochemical / genetical mechanisms determined by behavioural mechanisms [1] Libertini G (2012) Classification of Phenoptotic Phenomena. Biochem (Mosc) 77:707-15 3/20

4. Phenoptosis is unthinkable in terms of individual selection [1] (i.e., the famous Spencerian and then Darwinian “survival of the fittest” [2, 3]) ΔcS  P (1) [Δc = frequency variation between one generation and the next of a gene C that acts in the individual I; S = advantage / disadvantage for I caused by the gene C; P = reproductive value of I at the age when the gene manifests its action] On the contrary, phenoptosis is well explainable in terms of inclusive fitness [4-6] (of which individual selection and some types of group selection are particular or derived cases [1]). n Δc (SxPxrx) (2) x=1 [n = n. of individuals IX (I1, I2, I3, ..., In) genetically related to I1 for which the actions of the gene C have any effect; Sx = advantage / disadvantage for the individual IX; Px = reproductive value of an individual IX at the age when the gene C acts; rx = coefficient of relationship between individual IX and individual I1.] [1] Libertini G (2012) Classification of Phenoptotic Phenomena. Biochem (Mosc) 77:707-15 [2] Spencer H (1864) The Principles of Biology. Williams and Norgate, London [3] Darwin CR (1869) Origin of Species, 5th ed. John Murray, London [4] Hamilton WD (1964) The Genetical Evolution of Social Behaviour. J Theor Biol 7:1-52 [5] Hamilton WD (1970) Selfish and Spiteful Behaviour in an Evolutionary Model. Nature 228:1218-20 [6] Trivers RL (1971) The evolution of reciprocal altruism. Quart Rev Biol 46:35-57 4/20

5. Phenoptotic phenomena are well known and widespread in the living world [1]. Some examples: In the prokaryote world, there is the mass suicide of bacterial phytoplankton as defence against the propagation of phages [2] (direct optional phenoptosis). Likewise, phage infection activates the bacterial suicide in E. coli “thereby curtailing viral multiplication and protecting nearby E. coli from infection” [3]. phage infection single or mass bacterial suicide In the unicellular eukaryote world, in yeast, the suicide by apoptosis is activated in older individuals (i.e., cells of the mother lineage with more previous divisions) by the shortage of nutrients, and their cellular parts are useful to survive for other individuals [4](direct optional phenoptosis). shortage of nutrients suicide of “older” yeast individuals [1] Finch CE (1990) Longevity, Senescence, and the Genome. University of Chicago Press, Chicago [2] Lane N (2008) Marine microbiology: origins of death. Nature 453:583-5 [3] Raff MC (1998) Cell suicide for beginners. Nature 396:119-22 [4] Granot D, Levine A, Dor-Hefetz E (2003) Sugar-induced apoptosis in yeast cells. FEMS Yeast Res 4:7-13 5/20

6. For a lot of species(many plants, in particular monocarpic angiosperms, and animals, in particular Anguilliformes and Salmoniformes, some rodents and dasyurid marsupials, etc.), after reproduction there is rapid collapse of all vital functions and death (semelparity) [1](direct obligatory phenoptosis). Glycine max (soybeans) Antechinus stuartii Macleay's marsupial mouse) Phyllostachys bambusoides (giant or Japanese timber bamboo). Semelparity is not at all a synonym of short life. E.g., the giant timber bamboo blooms after 120 years and then dies [2]. Oncorhyncus nerka (Sockeye or Pacific salmon) [1] Finch CE (1990) Longevity, Senescence, and the Genome. University of Chicago Press, Chicago [2] Kawamura S (1927) On the periodical flowering of the bamboo. Jpn J Bot 3:335-49 6/20

7. Many types of insects, e.g., mayflies (Epheromoptera), in the adult phase are completely unable to feed because they have incomplete masticatory apparatus (aphagy) [1](direct obligatory phenoptosis). Other insects in the adult phase can live only a few weeks as they do not have mechanisms for renewing their anatomical structures. The housefly (Musca domestica) and the fruit fly (Drosophila melanogaster) are examples of these insects [1](direct obligatory phenoptosis). In some cases the survival of the offspring necessarily implies the death of the mother. For example, for the nematode Ascaris nigrovenosa the larvae pass through the mother's body (endotokic matricide) and for the beetle Micromalthus debilis the larvae feed on mother's body [1]. The sacrifice of these mothers is a type of direct obligatory phenoptosis. A mayfly Musca domestica Micromalthus debilis [1] Finch CE (1990) Longevity, Senescence, and the Genome. University of Chicago Press, Chicago 7/20

8. In other cases, the male dies shortly after mating with the female (e.g., male bees [1] and ants [2])(direct obligatory phenoptosis). In some cases, in order to procreate, the male faces a high risk of dying because at the moment of copulation the female tries to kill and devour him (e.g., the male of Mantis religiosa) [3](direct optional phenoptosis). Mantis religiosa (European mantis) A honeybee queen and a drone [1] Winston ML (1987) The Biology of the Honey Bee. Harvard Univ. Press, London [2] Holldobler B, Wilson EO (1990) The Ants. Springer-Verlag, Berlin-Heidelberg [3] Lawrence SE (1992). Sexual cannibalism in the praying mantid, Mantis religiosa: a field study. Animal Behaviour. 43(4):569-83 8/20

9. In many cases (e.g., in social vertebrates species), unselfish behaviours that are dangerous for one’s survival but increase the survival probabilities of others are common [1] (direct optional phenoptosis by behavioural mechanisms). For example, the predominant males of Papio cynocephalus (yellow baboon) [2] and of Papio ursinus (chacma baboon) [3] place themselves, with great individual risk, in the most exposed positions to defend from predators their herd. For various species of birds, there are the distraction behaviours shown by parents with great individual risk to save offspring threatened by predators [4, 5]. [1] Wilson EO (1975) Sociobiology, The New Synthesis. Harvard University Press, Cambridge [2] Altman SA, Altmann J (1970) Baboon ecology: African field research. Univ. of Chicago Press, Chicago [3] Hall KR (1960) Social vigilance behaviour in of the chacma baboon, Papio ursinus. Behaviour 16:261-94 [4] Brown RG (1962) The aggressive and distraction behaviour of the Western Sandpiper. Ibis 104:1-12 [5] Gramza AF (1967) Responses of brooding nighthawks to a disturbance stimulus. Auk 84(1):72-86 9/20

10. Since an individual may die only once, possible wrong deductions are that: • each species can present only a single phenoptotic phenomenon; • the species are divided between phenoptotic and non-phenoptotic ones. NO! These are erroneous ideas. Even the example of a single, well studied species (our species) shows their incorrectness. 10/20

11. Before birth, a fetus is eliminated in many cases (indirect phenoptosis by biochemical mechanisms)[1]. Somehow organism identifies defective fetuses or conditions that can cause problems and activates, as evolutionary physiologic solution, mechanisms of indirect phenoptosis. 1) The most common case is when there are abnormalities in the fetus: “nearly 70% of human conceptions do not survive to live birth … Overall, 50%–70% of specimens from sporadic spontaneous losses show some type of cytogenetic abnormality” [2] [1] Libertini G (2012) Classification of Phenoptotic Phenomena. Biochem (Mosc) 77:707-15 [2] Hyde KJ, Schust DJ (2015) Genetic Considerations in Recurrent Pregnancy Loss. Cold Spring Harb Perspect Med 5(3): a023119 11/20

12. 2) For our species, in the wild, it is difficult or even death-causing to rear two children at the same time. This motivates the phenomenon defined as the “vanishing twin” [1]: “... twins are conceived two to four times more often than they are born; in the majority of cases, the smaller of the two foetuses disappears by the third trimester and is apparently reabsorbed by the mother …” [2]. 3) Non-pathologic spontaneous miscarriages eliminate the fetuses with lesser antigen variability that have a likely reduced resistance to antigen mimicry of infective agents [3]. A greater pregnancy loss rate [4] and significantly fewer children [5] was the rule in the case of shared antigenic alleles. [1] Landy HJ, Keith LG (1998) The vanishing twin: a review. Human Reprod Update 4:177-83 [2] Hausfater G, Hrdy SB (1984) Infanticide: Comparative and evolutionary perspectives. Aldine, New York [3] Apanius V, et al. (1997) The nature of selection on the major histocompatibility complex. Crit Rev Immunol 17:179-224 [4] Ober C, et al. 1998) Human leukocyte antigen matching and fetal loss: results of a 10 year prospective study. Hum Reprod 13:33-8 [5] Ober C, van der Ven K (1997) Immunogenetics of reproduction: an overview. In: Olding LB, ed, Current Topics in Microbiology and Immunology. Springer-Verlag, Berlin 12/20

13. After birth, in primitive conditions, in many cases children are killed [1](indirect phenoptosis by behavioural mechanisms). The filial infanticide is documented for many human societies [2]. Among the motivations found in 60 different mostly primitive societies, we have [1]: Deformed or very ill newborn (21), Twin birth (14), Birth too soon or too many children (11), No male support (6), Mother unwed (14), Economic hardship (3). The newborn is killed usually by parents. It could be objected that these events are an expression of the primitive conditions of almost all the societies studied, but in modern societies a remarkable number of fetuses is eliminated by abortion for a series of analogous or identical reasons. [1] Hausfater G, Hrdy SB, eds, Infanticide: comparative and evolutionary perspectives. Aldine, New York [2] Scrimshaw SCM (1984) Infanticide in human populations: societal and individual concerns. In: Hausfater G, Hrdy SB, eds, Infanticide: … 13/20

14. In adulthood, there are numberless circumstances in which an individual may endanger or even sacrifice his / her life [1] … --- to save or try to save one or more genetically related individuals; --- … or also non-genetically related individuals; --- to defend his group from animals or from other groups; --- to attack other groups; --- to get or maintain a mate; … (direct optional phenoptosis) [1] Libertini G (2012) Classification of Phenoptotic Phenomena. Biochem (Mosc) 77:707-15 14/20

15. For the individuals not died by other events, there is the death due to the manifestations of aging (obligatory “slow phenoptosis” [1]). Subject of great discussion is the dilemma between two opposing interpretations of aging: 1) Non-programmed (non-adaptive) paradigm, which means that aging is seen as a failure of evolution and NOT as a phenoptotic phenomenon 2) Programmed (adaptive) paradigm, which means that aging is seen as an achievement of evolution and a phenoptotic phenomenon The first, or old, paradigm is still very widespread [2], but many facts and arguments are against it and in support of the second paradigm[3]. [1] Skulachev VP (2002) Programmed Death Phenomena: From Organelle to Organism. Ann N Y Acad Sci 959:214-37 [2] Olshansky SJ et al. (2002) Position statement on human aging. J Gerontol A Biol Sci Med Sci 57(8):B292-7 [3] Libertini G (2015) Non-programmed versus programmed aging paradigm. Curr Aging Sci 8:56-68 15/20

16. However, there is a general observation that supports the thesis that aging is an adaptive and programmed phenomenon. Aging is a phenomenon shown only by a minority of species. Most species do not show any age-related mortality increase, i.e., do not age, or show other types of life tables [1]. A semelparous species (Octopus maya, Mexican four-eyed octopus) An aging species (Panthera leo, lion) A non-aging species (Quercus rugosa, netleaf oak) to be continued … [1] Jones et al. (2014) Diversity of ageing across the tree of life. Nature 505:169-73 16/20

17. The life tables of living species show a wide variety of types [1]. Disregarding the species with constant mortality or the species that age, the other species have life tables that are surely genetically determined and favoured by natural selection. If the life tables of aging species (a minority) were not genetically determined and not favoured by natural selection, this would be a strange and unlikely exception in the living world. [1] Jones et al. (2014) Diversity of ageing across the tree of life. Nature 505:169-73 17/20

18. Furthermore, each individual of any eukaryotic multicellular species, our species included, originates from the evolution of monoclonal colonies of eukaryotic cells. In these colonies, only the cells of the germ line reproduce while for all the others there is always the availability to the sacrifice. Non-germ line cells die by one of several types of programmed cell death (PCD), e.g., apoptosis, keratinization and cell detachment of skin cells, etc. Each day, about 50 to 70 billion cells perish in the average adult because of PCD [1], i.e., about 700.000 cells per second. These cell deaths are not within the definition of phenoptosis but originates from a sort of phenoptotic pact between the cells of the original monoclonal colony. [1] Reed JC (1999) Dysregulation of Apoptosis in Cancer. J. Clin. Oncol. 17, 2941-53 18/20

19. Conclusion Phenoptotic phenomena are not a collection of rare and exceptional phenomena of marginal importance for the living beings and for evolution. On the contrary, phenoptotic phenomena are widespread and essential throughout the living world. Life tables and many characteristics of living beings may be incomprehensible if the concept of phenoptosis is neglected. For the mechanisms of supra-individual selection (kin and group selection, selective mechanisms in holobionts, selective mecha-nisms in ecosystems) it is essential to consider that, in evolutionary terms, life (survival of the fittest) and death (sacrifice of the individual or of an offspring) are closely interconnected. … to be continued

20. Conclusion (continued) The concept of phenoptosis must be considered an essential and pivotal idea in evolutionism. Survival of the fittest and phenoptotic death when it is useful Survival of the fittest (evolution of evolutionism) So, welcome in the phenoptotic world and thanks for your attention! This oral presentation is also available on my personal pages: www.r-site.org/ageing and on www.researchgate.net (e-mail: giacinto.libertini@tin.it)