Casz compositions and methods of use

Abstract

Provided are compositions and methods that include one or more of: (1) a “CasZ” protein (also referred to as a CasZ polypeptide), a nucleic acid encoding the CasZ protein, and/or a modified host cell comprising the CasZ protein (and/or a nucleic acid encoding the same); (2) a CasZ guide RNA that binds to and provides sequence specificity to the CasZ protein, a nucleic acid encoding the CasZ guide RNA, and/or a modified host cell comprising the CasZ guide RNA (and/or a nucleic acid encoding the same); and (3) a CasZ transactivating noncoding RNA (trancRNA) (referred to herein as a “CasZ trancRNA”), a nucleic acid encoding the CasZ trancRNA, and/or a modified host cell comprising the CasZ trancRNA (and/or a nucleic acid encoding the same).

1 - 86 . (canceled) 
     
     
         87 . A composition comprising a programmable nuclease having a length of no more than 900 amino acids, or a nucleic acid encoding the programmable nuclease, and a non-naturally occurring or engineered guide nucleic acid comprising a region that binds to the programmable nuclease and a guide sequence that is complementary to a target sequence of a target nucleic acid, or a nucleic acid encoding the guide nucleic acid. 
     
     
         88 . The composition of  claim 87 , further comprising a transactivating noncoding RNA. 
     
     
         89 . The composition of  claim 87 , further comprising the target nucleic acid. 
     
     
         90 . The composition of  claim 87 , wherein the target nucleic acid is single stranded DNA. 
     
     
         91 . The composition of  claim 89 , wherein the target nucleic acid is single stranded DNA. 
     
     
         92 . The composition of  claim 90 , wherein the target nucleic acid lacks a protospacer adjacent motif (PAM) sequence. 
     
     
         93 . The composition of  claim 91 , wherein the target nucleic acid lacks a PAM sequence. 
     
     
         94 . The composition of  claim 87 , wherein the target nucleic acid is double stranded DNA. 
     
     
         95 . The composition of  claim 89 , wherein the target nucleic acid is double stranded DNA. 
     
     
         96 . The composition of  claim 87 , wherein the target nucleic acid is a eukaryotic target DNA. 
     
     
         97 . The composition of  claim 89 , wherein the target nucleic acid is a prokaryotic target DNA. 
     
     
         98 . The composition of  claim 87 , wherein the target nucleic acid is a prokaryotic target DNA. 
     
     
         99 . The composition of  claim 87 , wherein the target nucleic acid is a viral target DNA 
     
     
         100 . The composition of  claim 89 , wherein the target nucleic acid is a viral target DNA. 
     
     
         101 . The composition of  claim 87 , further comprising a donor polynucleotide. 
     
     
         102 . The composition of  claim 87 , further comprising a cell. 
     
     
         103 . The composition of  claim 102 , wherein the cell comprises the programmable nuclease and the non-naturally occurring or engineered guide nucleic acid. 
     
     
         104 . The composition of  claim 102 , wherein the cell is a eukaryotic cell. 
     
     
         105 . The composition of  claim 87 , wherein the programmable nuclease comprises three partial RuvC domains. 
     
     
         106 . The composition of  claim 87 , wherein the programmable nuclease comprises RuvC-I, RuvC-II, and RuvC-III subdomains. 
     
     
         107 . The composition of  claim 87 , wherein the programmable nuclease is a CasZ protein. 
     
     
         108 . The composition of  claim 87 , wherein the programmable nuclease is selected from a group consisting of a CasZa protein, a CasZb protein, a CasZc protein, a CasZd protein, a CasZe protein, a CasZf protein, a CasZg protein, a CasZh protein, a CasZi protein, a CasZj protein, a CasZk protein, and a CasZl protein. 
     
     
         109 . The composition of  claim 87 , wherein the programmable nuclease is a variant programmable nuclease with reduced nuclease activity compared to a corresponding wild type programmable nuclease. 
     
     
         110 . The composition of  claim 87 , wherein the programmable nuclease is a dead programmable nuclease. 
     
     
         111 . The composition of  claim 87 , wherein the programmable nuclease is conjugated to a heterologous moiety. 
     
     
         112 . The composition of  claim 111 , wherein the heterologous moiety is a heterologous polypeptide. 
     
     
         113 . The composition of  claim 112 , wherein the heterologous polypeptide comprises a nuclear localization signal. 
     
     
         114 . The composition of  claim 87 , wherein the programmable nuclease has a length of from 350 to 900 amino acids. 
     
     
         115 . The composition of  claim 87 , wherein the programmable nuclease has a length of no more than 800 amino acids. 
     
     
         116 . The composition of  claim 87 , wherein the guide nucleic acid is a guide RNA.