{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/108686"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/108686","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Dynamic changes of TDP-43 in testicular and epididymal sperm","abstract":"TAR DNA-binding protein of 43 kDa (TDP-43) is a ubiquitously expressed and evolutionarily conserved protein. TDP-43 is a DNA/RNA binding protein with several functions such as gene transcription, mRNA splicing and stability, transposon silencing, and micro RNA biogenesis. TDP-43 is associated with many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Our lab previously demonstrated that sperm from infertile men contains aberrant forms of TDP-43. We also characterized TDP-43 expression in the mouse testis. However, little is known about the functions of TDP-43 during sperm epididymal maturation. One in six couples worldwide struggles with infertility with male factor accounting for half the cases, highlighting the importance of better understanding the mechanisms regulating spermatogenesis and sperm maturation. The goal of the present study was to evaluate the dynamic changes of TDP-43 from late spermiogenesis through sperm epididymal maturation. I hypothesized that TDP-43 is acquired by spermatozoa during epididymal maturation and undergoes post-translational modification during epididymal transit. In this study, I used three separate TDP-43 antibodies to detect whether TDP-43 was present in Stage VIII step 16 elongated spermatids using indirect immunofluorescence. My results indicated that TDP-43 is not present in step 16 elongated spermatids. At spermiation, spermatozoa are released into the lumen of the seminiferous tubule and are functionally immature; spermatozoa enter the epididymis where biochemical changes lead to sperm maturation and ability to swim. However, the processes of epididymal maturation are unknown; therefore, one goal of this study was to determine the localization and biochemical changes of TDP-43 in cauda epididymal spermatozoa. Indirect immunofluorescence using two TDP-43 antibodies showed that TDP-43 is localized in the acrosome and equatorial segment of caudal sperm. This suggests that TDP-43 is acquired during sperm epididymal maturation. Next, using western blot analyses, I showed that a predominant 55 kDa TDP-43 band was present in cauda epididymal sperm, whereas the testis protein extracts showed the monomeric form at 43 kDa. Immunoblot analysis of testis and epididymal extracts from 30-day old mice showed that the above 55 kDa is only weakly present in the epididymis at this age. This suggested that 1. The epididymal tissue expresses TDP-43, and 2. Sperm entry (which occurs at day 35 and onwards) causes increased expression of TDP-43. Use of pS409/410 and pS403/404 TDP-43 antibodies in immunoblots indicated that TDP-43 undergoes Serine phosphorylation resulting in the slower migrating 55 kDa epididymal sperm-specific TDP-43 species. Taken together, my results suggest that the epididymal maturation process adds TDP-43 to sperm. The presence of TDP-43 in the acrosome/equatorial segment of sperm is indicative of a role in sperm-zona or sperm-egg fusion. Alternatively, the RNA binding property of TDP-43 might serve the function of carrying RNAs into the egg. Future studies will test these hypotheses regarding TDP-43’s putative role in fertilization and/or early embryo development.","abstract_html":"TAR DNA-binding protein of 43 kDa (TDP-43) is a ubiquitously expressed and evolutionarily conserved protein. TDP-43 is a DNA/RNA binding protein with several functions such as gene transcription, mRNA splicing and stability, transposon silencing, and micro RNA biogenesis. TDP-43 is associated with many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Our lab previously demonstrated that sperm from infertile men contains aberrant forms of TDP-43. We also characterized TDP-43 expression in the mouse testis. However, little is known about the functions of TDP-43 during sperm epididymal maturation. One in six couples worldwide struggles with infertility with male factor accounting for half the cases, highlighting the importance of better understanding the mechanisms regulating spermatogenesis and sperm maturation. The goal of the present study was to evaluate the dynamic changes of TDP-43 from late spermiogenesis through sperm epididymal maturation. I hypothesized that TDP-43 is acquired by spermatozoa during epididymal maturation and undergoes post-translational modification during epididymal transit. In this study, I used three separate TDP-43 antibodies to detect whether TDP-43 was present in Stage VIII step 16 elongated spermatids using indirect immunofluorescence. My results indicated that TDP-43 is not present in step 16 elongated spermatids. At spermiation, spermatozoa are released into the lumen of the seminiferous tubule and are functionally immature; spermatozoa enter the epididymis where biochemical changes lead to sperm maturation and ability to swim. However, the processes of epididymal maturation are unknown; therefore, one goal of this study was to determine the localization and biochemical changes of TDP-43 in cauda epididymal spermatozoa. Indirect immunofluorescence using two TDP-43 antibodies showed that TDP-43 is localized in the acrosome and equatorial segment of caudal sperm. This suggests that TDP-43 is acquired during sperm epididymal maturation. Next, using western blot analyses, I showed that a predominant 55 kDa TDP-43 band was present in cauda epididymal sperm, whereas the testis protein extracts showed the monomeric form at 43 kDa. Immunoblot analysis of testis and epididymal extracts from 30-day old mice showed that the above 55 kDa is only weakly present in the epididymis at this age. This suggested that 1. The epididymal tissue expresses TDP-43, and 2. Sperm entry (which occurs at day 35 and onwards) causes increased expression of TDP-43. Use of pS409/410 and pS403/404 TDP-43 antibodies in immunoblots indicated that TDP-43 undergoes Serine phosphorylation resulting in the slower migrating 55 kDa epididymal sperm-specific TDP-43 species. Taken together, my results suggest that the epididymal maturation process adds TDP-43 to sperm. The presence of TDP-43 in the acrosome/equatorial segment of sperm is indicative of a role in sperm-zona or sperm-egg fusion. Alternatively, the RNA binding property of TDP-43 might serve the function of carrying RNAs into the egg. Future studies will test these hypotheses regarding TDP-43’s putative role in fertilization and/or early embryo development.","abstract_has_math":false,"creators":["Miller, Lyndzi Marie"],"institution":"University of Illinois at Urbana-Champaign","degree_name":"M.S.","degree_level":"Thesis","degree_discipline":"VMS - Comparative Biosciences","degree_department":null,"school":null,"contributors":["Reddi, Prabhakara P","Flaws, Jodi A","Nowak, Romana A"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2020,"date_issued":"2020-10-07T22:49:51Z","date_published":"2020-10-07T22:49:51Z","updated_at":"2026-07-22T22:24:48Z","subjects":["male reproduction","spermatogenesis","testis","epididymis","sperm","TDP-43"],"languages":["en"],"rights":["Copyright 2020 Lyndzi Miller"],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"http://hdl.handle.net/2142/108686","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Reddi, Prabhakara P","Flaws, Jodi A","Nowak, Romana A"]},{"key":"dc:creator","label":"Author","values":["Miller, Lyndzi Marie"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2020-10-07T22:49:51Z","2022-10-07T22:50:13Z","2020-07-16","2020-08"]},{"key":"dc:type","label":"Dc Type","values":["text","Thesis"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["VMS - Comparative Biosciences"]},{"key":"thesis:degree_level","label":"Degree Level","values":["Thesis"]},{"key":"thesis:degree_name","label":"Degree Name","values":["M.S."]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["University of Illinois at Urbana-Champaign"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["male reproduction","spermatogenesis","testis","epididymis","sperm","TDP-43"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["en"]},{"key":"dc:rights","label":"Dc Rights","values":["Copyright 2020 Lyndzi Miller"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["http://hdl.handle.net/2142/108686"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["TAR DNA-binding protein of 43 kDa (TDP-43) is a ubiquitously expressed and evolutionarily conserved protein. TDP-43 is a DNA/RNA binding protein with several functions such as gene transcription, mRNA splicing and stability, transposon silencing, and micro RNA biogenesis. TDP-43 is associated with many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Our lab previously demonstrated that sperm from infertile men contains aberrant forms of TDP-43. We also characterized TDP-43 expression in the mouse testis. However, little is known about the functions of TDP-43 during sperm epididymal maturation. One in six couples worldwide struggles with infertility with male factor accounting for half the cases, highlighting the importance of better understanding the mechanisms regulating spermatogenesis and sperm maturation. The goal of the present study was to evaluate the dynamic changes of TDP-43 from late spermiogenesis through sperm epididymal maturation. I hypothesized that TDP-43 is acquired by spermatozoa during epididymal maturation and undergoes post-translational modification during epididymal transit. In this study, I used three separate TDP-43 antibodies to detect whether TDP-43 was present in Stage VIII step 16 elongated spermatids using indirect immunofluorescence. My results indicated that TDP-43 is not present in step 16 elongated spermatids. At spermiation, spermatozoa are released into the lumen of the seminiferous tubule and are functionally immature; spermatozoa enter the epididymis where biochemical changes lead to sperm maturation and ability to swim. However, the processes of epididymal maturation are unknown; therefore, one goal of this study was to determine the localization and biochemical changes of TDP-43 in cauda epididymal spermatozoa. Indirect immunofluorescence using two TDP-43 antibodies showed that TDP-43 is localized in the acrosome and equatorial segment of caudal sperm. This suggests that TDP-43 is acquired during sperm epididymal maturation. Next, using western blot analyses, I showed that a predominant 55 kDa TDP-43 band was present in cauda epididymal sperm, whereas the testis protein extracts showed the monomeric form at 43 kDa. Immunoblot analysis of testis and epididymal extracts from 30-day old mice showed that the above 55 kDa is only weakly present in the epididymis at this age. This suggested that 1. The epididymal tissue expresses TDP-43, and 2. Sperm entry (which occurs at day 35 and onwards) causes increased expression of TDP-43. Use of pS409/410 and pS403/404 TDP-43 antibodies in immunoblots indicated that TDP-43 undergoes Serine phosphorylation resulting in the slower migrating 55 kDa epididymal sperm-specific TDP-43 species. Taken together, my results suggest that the epididymal maturation process adds TDP-43 to sperm. The presence of TDP-43 in the acrosome/equatorial segment of sperm is indicative of a role in sperm-zona or sperm-egg fusion. Alternatively, the RNA binding property of TDP-43 might serve the function of carrying RNAs into the egg. Future studies will test these hypotheses regarding TDP-43’s putative role in fertilization and/or early embryo development.","Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2022-08-01","The student, Lyndzi Miller, accepted the attached license on 2020-07-12 at 22:40.","The student, Lyndzi Miller, submitted this Thesis for approval on 2020-07-12 at 23:14.","This Thesis was approved for publication on 2020-07-16 at 18:35.","DSpace SAF Submission Ingestion Package generated from Vireo submission #15557 on 2020-10-02 at 15:50:12","Made available in DSpace on 2020-10-07T22:49:51Z (GMT). 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TDP-43 is a DNA/RNA binding protein with several functions such as gene transcription, mRNA splicing and stability, transposon silencing, and micro RNA biogenesis. TDP-43 is associated with many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Our lab previously demonstrated that sperm from infertile men contains aberrant forms of TDP-43. We also characterized TDP-43 expression in the mouse testis. However, little is known about the functions of TDP-43 during sperm epididymal maturation. One in six couples worldwide struggles with infertility with male factor accounting for half the cases, highlighting the importance of better understanding the mechanisms regulating spermatogenesis and sperm maturation. The goal of the present study was to evaluate the dynamic changes of TDP-43 from late spermiogenesis through sperm epididymal maturation. I hypothesized that TDP-43 is acquired by spermatozoa during epididymal maturation and undergoes post-translational modification during epididymal transit. In this study, I used three separate TDP-43 antibodies to detect whether TDP-43 was present in Stage VIII step 16 elongated spermatids using indirect immunofluorescence. My results indicated that TDP-43 is not present in step 16 elongated spermatids. At spermiation, spermatozoa are released into the lumen of the seminiferous tubule and are functionally immature; spermatozoa enter the epididymis where biochemical changes lead to sperm maturation and ability to swim. However, the processes of epididymal maturation are unknown; therefore, one goal of this study was to determine the localization and biochemical changes of TDP-43 in cauda epididymal spermatozoa. Indirect immunofluorescence using two TDP-43 antibodies showed that TDP-43 is localized in the acrosome and equatorial segment of caudal sperm. This suggests that TDP-43 is acquired during sperm epididymal maturation. Next, using western blot analyses, I showed that a predominant 55 kDa TDP-43 band was present in cauda epididymal sperm, whereas the testis protein extracts showed the monomeric form at 43 kDa. Immunoblot analysis of testis and epididymal extracts from 30-day old mice showed that the above 55 kDa is only weakly present in the epididymis at this age. This suggested that 1. The epididymal tissue expresses TDP-43, and 2. Sperm entry (which occurs at day 35 and onwards) causes increased expression of TDP-43. Use of pS409/410 and pS403/404 TDP-43 antibodies in immunoblots indicated that TDP-43 undergoes Serine phosphorylation resulting in the slower migrating 55 kDa epididymal sperm-specific TDP-43 species. Taken together, my results suggest that the epididymal maturation process adds TDP-43 to sperm. The presence of TDP-43 in the acrosome/equatorial segment of sperm is indicative of a role in sperm-zona or sperm-egg fusion. Alternatively, the RNA binding property of TDP-43 might serve the function of carrying RNAs into the egg. Future studies will test these hypotheses regarding TDP-43’s putative role in fertilization and/or early embryo development.","Submission published under a 24 month embargo labeled 'Closed Access', the embargo will last until 2022-08-01","The student, Lyndzi Miller, accepted the attached license on 2020-07-12 at 22:40.","The student, Lyndzi Miller, submitted this Thesis for approval on 2020-07-12 at 23:14.","This Thesis was approved for publication on 2020-07-16 at 18:35.","DSpace SAF Submission Ingestion Package generated from Vireo submission #15557 on 2020-10-02 at 15:50:12","Made available in DSpace on 2020-10-07T22:49:51Z (GMT). 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